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- 1. Acrosome is a specialized form of the golgi that migrates to the tip
- 2. The mid-piece houses mitochondria that produce GTP needed by the dynein for flagellar movement
- 3. Sperm are produced throughout the life of the male
- Before Puberty:
- 1. Primary oocytes are arrested in prophase of meiosis I
- 2. The primary oocyte grows and ovarian stromal cells form a single layer of flattened follicular epithelial cells
- 3. A specialized coat called the zona pellucida forms
- 4. Cortical granules are placed at the periphery of the oocyte... it is now a primordial follicle
- After Puberty:
- 5. Follicular epithelial cells become columnar, granulosa cells forming a primary follicle
- 6. In response to FSH, follicle cells secrete estrogen
- 7. The follicle that has the most FSH receptors will grow the most and will become and antral follicle and eventually a mature Graafian follicle
- 8. The oocyte completes Meiosis I to become a secondary oocyte and a first polar body
- 9. Meiosis II begins but is arrested in metaphase II
- 10. Only if it is fertilized, meiosis II is completed and the second polar body appears on the side of the mature oocyte
The HPO Axis
- 1. Hypothalamus makes Gonadotropin releasing hormone (GnRH)
- 2. GnRH acts on the anterior pituitary and stimulates it to make FSH and LH
- 3. FSH and LH act on the ovary:
- - FSH stimulates the ovarian follicles to grow, differentiate, and produce estrogem
- -LH stimulates ovulation, corpus luteum formation and progesterone production
- 4. Estrogen stimulates growth and differnetiation of the uterus, vagina, oviduct and the mammary glands
- 5. Progestin stimulates the build up of the endometrium during the luteal phase
Primary Lymphoid Organs
- 1. Thymus:
- -has CT capsule that branches into septa on the inside of the thymus
- -has epithelioreticular cells that serve as a support; also create the blood-thymus barrier; and these cells produce thymic hormones
- -Cortex: high concentration of immature thymocytes; macrophages are present and appear white on the slide
- -Inner medulla: lighter branched structure with fewer thymocytes; stimulate "self" expression and only 5-10% of T-cells pass this test; Hassall's Corpuscles are found here
- -cortico-medullary boundary is characterized by high endothelial venules that have a thick endothelium where t-cells can exit the thymus and enter the periphery
Bone Marrow: Immunocompetent B-cells are made here
Common Features of Secondary Lymphoid Organs
1. Nodular tissue: compact well circumscribed collections of lymphoid tissue, reticular cells and their associated fibers; mostly B-cells; can be primary (netwrok of reticular fibers and small, tightly packed lymphocytes) or secondary (stimulated cells in the germinal centers responding to antigen and proliferating B cells)
- 2. Diffuse Tissue: between the nodules and the connective tissues; mostly T-cells and macrophages supported by reticular cells
- 3. All secondary lymphoid organs are defined based on whether they are encapsulated or not
Unencapsulated Secondary Lyphoid Tissue
MALT (Mucosa Associated Lymphoid Tissue)
- 1. Tonsils: masses of nodular and diffuse tissue with crypts to increase surface area; three types: palantine (paired in the back of the throat; stratified squamous); lingual (multiple on the back of toungue; stratified squamous); pharygeal (single, with pseudostratified ciliated columnar)
- 2. Peyer's Patches: nodules with intervening diffuse tissue in the mucosa of the ileum; have specialized M-cells that are surrounding lymphocytes and sample antigen in the lumen and transfer it to them
- 3. Appendix: rich in nodular and diffuse lymphoid tissue and has M-cells
Encapsulated Secondary Lymphoid Tissue
- 1. Lymph Node
- -bean shaped organ surrounded by a dense tissue capsule and supported by reticular fibers that are continuous with the capsule
- -cortex is made up of nodules and diffuse tissue that extends into the medulla to form loose medullary cords that are surrounded by medullary sinuses (loose vessels of reticular fibers)
- -fluid enters the capsule through the afferent lymphatic and passes through the subcapsular sinus; these sinuses enter the medullary sinuses which are high endothelial venules; then exits through an efferent lymphatic called the hilus
- -the paracortex is right below the cortex and is a t-cell rich area
- 2. Spleen:
- -covered by mesothelium and an underlying thick capsule; trabecular beams penetrate the spleen
- -no cortex or medulla
- -white pulp: lymphoid tissue, both nodular and diffuse organized around arteries
- -red pulp: organized around venous sinuses to increase contact between red blood cells and the macrophages
- -open circulation: sheathed capillaries pass through the white pulp then empty into the red pulp; there is only one hilus but many afferent vessels so fluid slows down
What is the innate immunity and what does it include?
- -a variety of machinsms that can prevent of eliminate infection
- -present at all times; does not increase with repeated exposure to a pathogen
- -physical barriers: skin, mucosous membranes
- -biologically active substances: lysozyme; cytokines, anti-microbial proteins (ont he skin) and activation of complement
- -cellular: natural killer cells and phagocytes (macrophages and neutrophils)
Cells of the innate immune system
- 1. Macrophage: phagocytic leukocytes found in almost all tissues; mononuclear and derived from blood monocytes
- 2.Neutrophils: phagocytic leukocytes that are important for engulfing and destroying extracellular pathogens, primarily bacteria; also known as polymorphonucelar leukocytes (PMNs) because of their multi-lobed nucelus
- 3. Eosiniphils: leuokcytes that kill antibody covered parasites; usually by releasing substances
- 4. Mast cell: congregate in tissues and release granule of histimine when activated; major role in allergci response
- 5. Basophils: circulate in blood and thought to have similar function to Mast cells
- 6. Natural Killer cells: derived from the lymphoid progenitor cells; destroy virus-infected and tumor cells without prior stimulation
What are PRRs and PAMPs?
PRR: Pattern recognition receptors; receptors for pathogens on the cells of the innate immune system; have evolved to recognize general features of pathogens
PAMP: a microbial product recognized by the PRR; usually features of the cell that are vital for function so they do not eveolve away
What is the Adpative/Acquired Immunity?
- -host defenses mediated by the response of antigen-specific lymphocytes
- -requires sensitization my antigen; response in antigen-specific; results in immunological memory
- -capable of responding to virtually any foreign molecule
- -can be classified into humoral (B-cell) or cell mediated (T-cell)
What is the general differnce between humoral immunity and cell-mediated immunity?
1. Humoral: mediated by antigen-specific antibodies produced by activated B-lymphocytes (plasma cells); the reaction occurs in the blood; can be transferred to immune naive individuals through teh serum
2. Cell-mediated: primarily involved antigen-specific T-cells; actualy t-cells must be transfered to immune niave individuals; helper T-cells function by producing cytokines while cytotoxic T-cells recognize and kill cells infected with a pathogen
What is Clonal Selection in B-cells?
- -B-cells have membrane bound antibody that is specific for a single anitgen (the immune system is comprised of billions of different lymphocytes each with their own specificity)
- -recognition of the antigen causes the cell to undergo proliferation
- -some cells will become memory cells and the rest of the cells will become effector cells; in the case of B-cells, the effector cells are plasma cells that secrete antigen-specific antibodies
Structure of the mitochondria
What is significant about mitochondrial DNA?
- double-stranded, circular DNA that is maternally transmitted
- up to 10,000 copies per cell
- encodes rRNAs, tRNAs and 13 of the proteins needed for oxidative phosphorylation
- has its own system for replication and transcription
Mitochondrial protein import
- the vast majority of proteins in the mitochondria are synthesized in the nucelus and imported into the organelle
- some proteins bound for the mitochondria contain a presequence/matrix targeting sequence on their N-terminus that directs them to the mitochondria; other proteins for the mitochondria will have internal coding information that directs them to the correct compartment of membrane of the mitochondria
- the process is facilited by translocases of the outer membrane (TOMs) and translocases of the inner membrane (TIMs)different pathways are used for proetins with the presequence and for those with internal coding information
- the process requires energy
- the proteins are folded when they get to the mitochondria and they spontaneously unfold and they pass through the transport proteins in the membranes
- -can make 35 moles of ATP per mole of glucose; we cannot use dietary ATP; every cell must be able to make its own to be used for energy
- -electrons move through a series of oxidation/reduction reactions where they are transfered from reduced, low reduction potential molecules to oxidized, high reduction potentail molecules; this releases free energy which can be used to make ATP or be released as heat
- 1. Respiratory chain
- -Complex I: oxidized NADH and transfers a pari of electron to coenzyme Q
- -Complex II: a group of 3 dehydrogenases that transfer a pari of electrons from succinate, acetyl CoA, or glycerol phosphate to coenzyme Q
- -Coenzyme Q: shuttles the electrons it has picked up from Complex I and/or II and transfers them to Complex III
- -Complex III: accepts a pair of electrons from Coenzyme Q and transfers one to cytochrome C
- -Cytochrome C: passes that electron on to Complex IV
- -Complex IV: transfers the electron from cytochrome C to the terminal electron acceptor, oxygen
2. Creation of an electrocehmical transmembrane potential: as ox/red reaction occur at complexes I, III and IV, protons are moved from the inside of the mitochondria to the intermembrane space
3. ATP Synthase (AKA Complex V) makes ATP using the proton motive force across the inner mitochondrial membrane
What are the prosthetic groups associated with the Respiratory Chain?
- NADH: coenzyme that is the electron donor at the beginning of the chain
- Complex I: FMN, flavin mononucleotide, derived from riboflavin
- Complex II: FAD, flavin adenine dinucleotide, derived from riboflavin
- Coenzyme Q: is itself a prosthetic group, AKA Ubiquinone, has an isoprenoid side chain that can be reduced to carry the electrons to the next complex
- Cytochrome C, Complex III and IV: heme, note that these cannot serve as oxygen carriers because of the coordination position of the iron
Inhibitors of the Respiratory Chain
- 1. Rotenone: commonly used insecticide that binds to and inhibits Complex I
- 2. Antimycin: inhibits Complex III
- 3. Cyaninde and Azide: bind to the oxidized form of cytochrome a3 so they inhibit Complex IV
Structure of ATP Synthase
- F1: on the matrix side of the IMM, has the catalytic activity
- F0: spans the IMM and forms a channel for protons
What are the uncouplers of the respiratory chain?
FCCP, dinitrophenols (small molecules), and UCP1 protein
What is the relationship between obesity and uncoupling of the respiratory chain?
- Brown fat contains UCP1 protein which is an uncoupler
- Uncoupling forces mitochondria to burn calories at a maximum rate without doing sufficient work (no ATP is formed)
What is ANT?
- Adenine Nucleotide Translocase: transports newly synthesized ATP from the mitochondrial matrix into the cytosol and ADP from the cytosol into the mitochondria
- it is inhibited by atractyloside
What is MPT?
- Mitochondrial Permeability Transition: loss of the inner mitochodrial membrane impermeability caused by extended opening of the ANT due to accumulation of calcium in the matrix
- Calcium-dependent binding of the matrix protein, cyclophilin D to ANT; can be inhibited by using cyclosporin A which inhibits cyclophilin D
- Results in the cessation of ATP synthesis, swelling of the matrix which disrupts the outer membrane, and release of apoptotic factors
- Clinical Correlate: this occurs in cardiac cells during an MI and in neuronal cells during a stroke
What is the CDR region of an antibody?
- Complenetarity Determining Regions: the region of the antibody with the greatest amino acid variability
- -often the part of the antibody that makes contact witht he antigen
What are the immunoglobulin isotypes?
-The subclasses are defined by having variable heavy-chains; the variability and change in the effect of these isotypes is determined by the Fc region
Where are the different immunoglobulin isotypes found?
- IgG: most common; high concentration in the serum and it diffuses to all tissues; can cross the placenta giving the fetus enough immunity for a couple of months until it can make its own
- IgM: found as a pentamer linked together with a J chain that is found primarily in the blood stream
- IgE: associated with the skin and the nucosal surfaces
- Monomeric IgA: found in the serum
- Dimeric/secretory IgA: on mucosal surfaces and in secertions such as breast milk
Function of antibody?
- 1. Neutralization of a virus or toxin: addition of antibody to the surface of a virus or toxin or even bacteria makes it unable to enter a cell and cause damage; usually mediated by IgG and IgA
- 2. Opsonization: coating a bacteria in IgG helps phagocytic cells to eat it because neutrophils and macrophages have Fc receptors that recognize and bind it
- 3. Antibody-dependent cell-mediated cytotoxicity (ADCC): IgG binds the surface of the target cell (usually one that has been infected with a virus or in cancerous); natural killer cells have Fc Receptors on their surface that recognize this antibody covered cell and induce it to undergo apoptosis
- 4. Mast Cell degranulation: Mast cells have Fc receptors on their surface that bind IgE before it binds to antigen; when the mast cell bound IgE binds an antigen, the cells release various substances, including histamine; this reaction is important for allergies and for protection against parasites
- 5. Complement system activation: IgG and IgM activate the complement systme which is a seriosu of host protein that mediates defense against extracellular pathogens, especially bacteria; this amplifies the inflammatory response and releases chemotactic factors and punctures holes in the bacterial cell walls and membranes
What is transformation?
- -the introduction of naked, or free DNA from the environement into a bacterial cell
- -a cell must be competent to take up the DNA from the environment which is determined by the presence or absence of specific proteins; a cell can acquire comptency or be naturally competent
What is transduction?
- the transfer of DNA from one bacteria to another by a bacteriophage vector
- Generalized transduction: the transducing virus packages random fragments of host DNA
- Specialized transduction: the viruses pick up genes that lie near the site of prophage integration
- the process is DNase immune because the DNA is protected by the viral capsid
What is conjugation?
- the direct transfer of DNA from one bacteriums to another
- bacterial cells capable of conjugation carry special genes that code for the sex pilus and other things necessary for the transfer of DNA; usually on a conjugal plasmid
- the transferable element is sent to the recipient bacteria as a single strand, so the donor keeps one copy
- often genes are on a circular plasmid that can replicate itself
- the newly acquired genes can either stay as a plasmid and get passed on to all duaghter cells or it can incorporate into the genome of the bacteria through homologous recombination (requires RecA)
What is a plasmid and how is it replicated?
- a non-essential, extra-chromosomeal DNA replicon that is usually circular
- they can: 1. carry genes necessary for pathogeneisis, 2. carry genes to confer drug resitance and 3. be spread among bacteria
- they always have a ori site that is specific tot he plasmid
- usually have encode one or more special proteins called rep that are requires for plasmid replication; usually help the host DNA polymerase recognize the ori
- plasmid segregation is highly regulated to make sure that each daughter cell gets a copy
What is plasmid copy number and compatibility?
- each particular plasmid has a characteristic copy number at which it is typically found within a cell; copy number is controlled by plasmid genes
- compatibility refers to if two different plasmids can both be in a cell at the same time; usually dependent upon whether or not they have the same oriR-rep system
- occurs in the ampulla
- sperm swim through the cervical canal and are drawn up through the uterus to the fallopian tubes by muscular contractions and ciliary action; the process is 2/3 work of the female and only 1/3 the male
- the process begins with contact between the sperm and the secondary oocyte and ends with comingling of the maternal and paternal chromosomes at metaphase of hte first mitotic division approx. 24 hours later
What do the ovaries do?
- produce estrogen and progesterone which are respinsible for female secondary sex characteristics and regualtion of pregnancy
- produce and maintain the oocytes
Important things to know about the mentrual cycle
- 1. Menstrual phase
- 2. Proliferative Phase: estrogen levels rise and stimulate regrowth of the functional layer of the endometrium
- 3. Secretory Phase: estrogen levels peak, then fall and progesterone levels increase; corpus luteum makes progesterone which stimulates the glandular epithelium to secrete mucous fluid in preparation for a possible pregnancy; if fertilization occurs, the blasotcyst implants on day 20 and hCG from the syncitiotrophoblast prevents the corpus luteum from degenerating so estrogen and progesterone are continued to be released; if no fertilization occurs, the corpus luteum degenerates, hormone levels fall and ischemic phase happens
- 4. Ischemic Phase: progesterone and estrogen levels fall rapidly and the functional endometrium becomes ischemic
Important things to know about ovulation
- when estrogen levels from the secondary follicle are high enough a surge of LH is released from the AP which triggers ovulation 12-24 hours later
- it also stimulates the primary oocyte within the secondary follicle to complete MI and arrest in metaphase of MII as a secondary oocyte now called a mature or Graafian follicle
- at ovulation the oocyte and some of the cumulus cells are expelled by fluid pressure and contraction of the smooth muscle in the theca externa, under the influence of prostaglandins
- the ovulated secondary oocyte is surrounded by the zona pellucida and one or more layers of follicular cells
What is the deal with the corpus luteum?
- After ovulation the follicle collapses to form the CL which secretes progesterone
- The progesterone prepares the endometrium for implantation of the zygote
- If fertilization occurs, hCG is secreted from the blastocyst which prevents the degeneration fo teh CL
- If there is no fertilization, the CL becomes the corpus albicans, a patch of white scar tissue and progesterone levels fall abruptly and the endometrium goes ischemic
Important things to know about sperm production
- occurs throughout reproductive life, from puberty until death
- sperm counts lower than 10 million/mL and motility levels below 20% are associated with infertility
- sperm must undergo capacitation before they can fertilize the oocyte; their glycoprotein coat and seminal proteins are removed from the acrosome surface; makes them more active and motile
- Then they must undergo the acrosome reaction; angiotensin converting enzyme (ACE) in the acrosome is needed to induce this
What is the acrosome reaction?
- The sperm contacts the oocyte and moves throught he follicular cells and is received by the zona pellucida protein ZP3 (this protein accelerates the reactiona nd is unique among species)
- The acrosome membrane of the sperm perforates due to the ACE and hyaluronidase and proteases from the acrosome break downt he zona pellucida so the sperm can enter
What is the cortical reaction?
- initiated at the "Eureka Moment" when the sperm contacts the oocyte membrane
- The plasma membrane of the sperm and oocyte fuse and the head and tail of the sperm enter the oocyte
- Cortical granule contents are released from the oocyte and the zona pellucida is modified through proteolysis and protein cross-linking to make it impenetrable to other sperm; called the block to polyspermy; assisted by a calcium pulse across the oocyte membrane
- The oocyte finishes MII and becomes a mature oocyte
Formation of the pronuclei
- The oocyte chromosomes condense and from a pronucleus while the newly arrived spem nucleus enlarges and forms the male pronucleus
- The chromosomes replicate
- Pronuclear membranes break down, chromosomes condense and line up for the first metaphase
- Fertilization is now over and a zygote is formed; it is unicellular and diploid
What is the oriT site?
- An origin of transfer
- specific sequence on the plasmid that is nicked by an endonuclease and the nicked strand is transferred to a recipient cell
What is the tra operon?
- encodes many genes and all are involved in conjugation
- some of the proteins: ones involved in synthesis of the sex pili, DNA transfer protein, pilin, etc.
What is the TraJ gene?
- a positive regulator of the tra operon encoded on the F plasmid
- there was a negative regulator in the operon but an insertion mutation has rendered it non-functional; therefore the tra operon of F is always expressed at high levels
What is surface exclusion?
- prevention of a cell that carries a conjugative plasmid from inheriting additional related conjugative plasmids
- proteins encoded on the F plasmid modify the bacterial cell surface to block the ability of the sex pilus encoded by that same plasmid to attach to the cell
What is zygotic induction?
- when DNA is transfered to a new cell through conjugation, it is stripped of associated proteins including repressors
- therefore, a gene on a newly inherited piece of DNA that was repressed in the donor because of a bound repressor protein will become expressed int he recipient
- most tra operons code for a repressor but it takes time before enough reoressor builds up to tirn off the tra genes
- this results in the rapid spread of such plasmids and their genes throughout a population
What is an Hfr strain?
- A bacterial cell in which the conjugal plasmid has become part of the chromosome
- This occurs because the F plasmid has many insertion sequences (IS) that recombine to random homologous sequences in the hist chromosome
- This allows parts of the host genome that are next to the plasmid genes to be transfered through conjugation as well because the tra genes are still expressed
What is an insertion sequence?
- small units of DNA which encode only functions involved in insertion events, usually justa transposase
- they are everywhere and can transpose from one site in a chromosome or plasmid to a second site on the same genome or any other DNA molecule present in the cell
- they have short, perfect, terminal repeats
What are transposons?
- mobile genetic elements containing additionsal genes unrelated to transposition
- Composite Transposons: a gene flanked by 2 IS sequences; everything betweent he IS sequences can be transposed or the ends can transpose independently
- Simple or Noncomposite Transposons: structurally similar to IS sequences but they carry a singel drug resistance gene, a transposase gene and a resolvase gene between the short terminal repeats; must always transpose as a unit
How is transposition regulated?
- Transposase acts as its own repressor protein
- it binds tot he promoter region to prevent over-production of the transposase
What is replicative transposition?
- a replicated copy of the transposon is inserted into a new site
- Steps include: 1. Copying the transposon 2. Co-integrate formation 3. Resolution of the co-integrate by Tn-encoded resolvase (more efficient) protein or host recombination enzymes such as recA
- the target site is duplicated, but this is usually restored when teh transposon leaves that site
What is significat about the R100 plasmid?
it carries several drug resistance genes that it probably accumulated as more and more Tns hopped onto the plasmid
What is V(D)J Recombination?
- the recombining of Ig and TCR genes ina variety of different ways to produce the diversity of receptors and antibodies present in the immune system
- Light Chain: has two isotypes kappa and lambda; the variable region os coded for by different combinations of the V and J regions; this VJ segment then hooks up with a constant region domain associated with that isotype
- Heavy Chains: variable region is coded for by the V, D and J regions in different combination; then it hooks up with the C region genes and first IgM and IgD are made because their C regions come first in the gene
What is RAG-1 and RAG-2?
- Recombination-activating genes-1 and -2
- express exclusively in developing lymphocytes and are required for the initial cutting of the Ig or TCR genes
- if a person is defective in one or both of these genes, they have sever combined immune deficiency (SCID) and no B or T cels are produced
- Note: it is DNA tha tis cut and ligated together
What are the four mechanisms we use to generate antibody diversity?
- Germline Diversity: refers to the fact that we each have multiple differnet V, D, and J gene segments and multiple alleles for each of these genes
- Combinatorial Diversity: refers to all of the possible cobinations of V,D, and J genes that can be produced
- Junctional Diversity: one of the most important parts in generating diversity; before V and J chains attach to eachother, an exonuclease randomly chews back amino acids and TdT randomly adds entirely new nucelotides to the ends of their segments
- Somatic Hypermutation (SHM): refers to random point mutations that occur in the variable regions of the immunoglobulin during clonal selection; immunoglobulins with higher affitiny for antigen are then selected for; this accounts for a more effective secondary immune response; this only occurs in B cells, not T cells
What is Class Switching?
- occurs in secondary lymphoid tissue after exposure to antigen
- Class switch recombination means that during clonal selection, different isotypes of the immunoglobulins can be made by rearranging the VDJ sequence and moving it next to the constant regions for IgG, IgE, and IgA
What is AID protein for?
- activation-induced cytidine deaminase
- enzyme required for both CSR and SMH
- patients with a deficiency in this gene have Hyper-IgM Syndrome-2
What are the four different themes for viral genomes?
- Polyprotein: genome is an mRNA with a poly-A tail that is translated into a polyprotein that is later cleaved
- Segmented Genome: genome is packaged into the virus as pieces of DNA that is transcribed and translated into seperate proteins
- Nested mRNAs: genome is an mRNA with a poly-A tail but the genome is alternatively translated into different mRNAs that become different proteins
- Differential splicing: genome is an mRNA that is spliced before being translated into a protein
How can you classify a virus based on early events?
What is a porphyrin?
- important prosthetic groups in biochemistry
- it is a ring structure made of four pyrroles; there are 8 substituent groups
- Type I means that all the groups are in the same positions
- Type III means that the orientation of the D ring is reversed (like in heme)
- Porphyrinogens are the reduced form and can be toxic if they accumulate
- Step 1: Succinyl CoA + Glycine --> delta-aminolevulinic acid (delta- ALA); occurs in the mitochondria
- Step 2: delta-ALA + delta-ALA--> porphobilinogen; occurs in the cytoplasm; the enzyme that does this step, delta-ALA dehyratase, needs zinc and is a target in lead poisoning
- Step 3: 4 x porphobilinogen--> uroporphyrinogen; it is type III; the orientation is dependent upon the cosynthase; deamination occurs; still in the cytoplasm
- Step 4: Uroporphyrinogen - 4 CO2 --> coproporphyrinogen; this makes the molecule more hydrophobic so it wil be happy in its pocket on the hemoglobin molecule; this moves back into the mitochondria
- Step 5: Decarboxylation of coproporphyrinogen type III--> protoporphyrinogen type III
- Step 6: Oxidation of protoporphyrinogen--> protoporphyrin
- Step 7: Protoporphyrin binds iron--> heme
Which intermediates in heme synthesis can become oxidized and what is the result of that?
- uroporphyrinogen and coproporphyrinogen and colorless but can undergo oxidation spontaneously to make uroporphyrin (type I or II) amd coproporphyrin (type I or III)
- if the colorless molecules accumulate they can become oxidized and colored and stain tissues
- oxidized portphyrins can also lead to photo-oxidative damage to skin due to photosensitivity
What is the regulating step in heme synthesis and how is it regulated?
- The first step is the regulated step and it is catalyzed by ALAS-2 in erythroid cells and by ALAS-1 in all other cells
- It is inhibited by heme, the end product
- Regulation of ALAS-1: inhibition of gene transcription through destabilizing mRNA; inhibition of transport of the enzyme into mitochondria; allosteric inhibition in the mitochondria; barbiturates increase heme synthesis
- Regulation of ALAS-2: iron increases translation of the enzyme by displacing the inhibitor; iron increases ferrochelatase activity; heme prevents ALAS-2 from being transported into the mitochondria
What are porphyrias?
- deficiencies of the heme synthetic enzymes
- associated with abnormally high levels of ALAS activity and an overporduction of the intermediates that are before the slow/deficient step
- may be heriditary or acquired
- may be erythropoietic or hepatic
- testing included urinalysis for PBG; treatment includes reducing ALAS activity (give hemin, glucose, and eliminate barbiturates)
What is Acute intermittent porphyria?
- reduced activity of PBG deaminase so PBG and delta-ALA accumulate
- there is no photosensitivity
- may be triggered by barbiturates or other drugs metabolized by the cytochrome P450 enzyme system
- Step 1: heme/hemoglobin + O2--> biliverdin; catalyzed by a mixed function oxidase, cytochrome P450; it is an intensely green compound
- Step 2: biliverdin is reduced to bilirubin, a yellow-brown compound; accumulation of this is jaundice
- Step 3: insoluble bilirubin is bound to albumin and transported to the liver
- Step 4: conjugation of bilirubin to glucuronic acid--> excretion in bile to the small intestine
- Step 5: bacteria in the small intestine carry out further metabolic reaction to make urobilin, which colors the urine yellow, and stercobilin, which colors the feces brown
Definition of a lipid
- Hydrophobic organic molecules that can be extracted from cells using non-polar solvents
- Associate with eachother to maximize exclusion of water
What are the two classes of lipids?
- 1. Fatty acids: include neutral fat (TAGs) and a variety of charged and uncharged membrane lipids; long chain of carbon and hydrogen with a terminal carboxyl group
- 2. Isoprenoids: derived from isperene units, and are oftern mulit-ring organic molecules; includes cholesterol, steroid hormones, and the fat-soluble vitamins
Fatty acid nomenclature
- carbons are numbers from the carboxyl carbon (1) or from the terminal carbon (omega)
- saturated FAs have no double bonds
- unsaturated FAs have one or more double bonds; if it has multiple double bonds it is refered to as polyunsaturated fatty acids (PUFAs)
- # of carbons:#of double bonds(delta)locations of double bondsomega number = number of carbons-the highest number carbon of double bond
- at neutral pH, they are amphipathic and associate with sodium ions; then called a "sodium ____ate"
What is TAG?
- major form of storage lipid and the bulk of dietary lipid
- composed of three fatty acids in ester linkage to the three hydroxyl groups on glycerol
- the 1 and 3 positions are symmetric
- no longer carries the negative charge and it is completely hydrophobic
- nomenclature: change the FA suffix from "ic acid" to "oyl" and list numerically in front of glycerol
What is emulsification of fat?
- break up fat droplets that arrive in the intestine in order to increase their surface area
- achieved by: body temperature helps liquify them; peristalsis helps physically crush fat droplets; the gall bladder releases bile salts
- bile salts coat the surface of fat droplets and allow them to be water-soluble
- emulsification accelerates as TAGs are borken down and more surface area is created
What is lipolysis?
- hydrolytic cleavage of TAGs to form 2 FAs and 2-MAG
- catalyzed by pancreatic lipase
- the 2-MAG is sufficiently soluble because of the 2 OH groups
Absorption of fat into intestinal mucosal cells
- glycerol, 2-MAG, and free fatty acids wil inter the cells by simple diffusion becuase they are water-soluble enough
- long-chain fatty acids and 2-MAG are more insoluble and therefore cannot hang out in the lumen for very long
- they self-aggregate, with bile salts to form micelles at the critical micelle concentration (CMC)
- as more fatty acids and 2-MAGS enter the intestinal mucosa cells, micelles begin to break down, making more monomers available for absorption
How do intestinal epithelial cells maintain the concentration gradients of lipids so that they continue to enter the cell by diffusion?
- the cells resynthesize TAGs
- Step 1: the fatty acid is activated in the cytoplasm to form a fatty acyl coenzyme A derivative; this created a thioester bond which is high energy; uses ATP and releases free phosphates which must be used by inorganic pyrophosphatase to shift the equilibrium forward
- Step 2: two activated FAs are the added to the 2-MAG; the 2 position is the same as dietary but the 1 and 3 positions are radnomized
What is steatorrhea?
- lipid in the stool
- can be caused by a biliary obstruction, pancreatic disease or ciliac disease (low surface area for absorption)
What is a chylomicron?
- large lipoprotein complexes whos emajor function is to transport dietary TAGs through the blood to other tissues
- TAGs and cholesterol esters are resynthesized inside the intestinal epithelial cell in the ER membrane
- once enough is built up, they bud off into the lumen of the ER as a membrane bound lipid droplet
- next, apoprotein B-48 is added, forming nascent chylomicron
- they are secreted in vesicles and the nascent chylomicrons enter the lymphatic system where apoproteins C-II and E are hyjacked from HDL to form a mature chylomicron
What is the function of Apo C-II?
- activates Lipoprotein Lipase (LPL) on the surface of blood vessels
- LPL hydrolyses TAGs completely to FAs and glycerol so it can be absorbed by the surrounding cells
What is the fate of chylomicron remnants?
- Apo E and Apo B-48 bind the LDL receptor-related protein (LRP) or remnant receptor on the surface of the liver
- the remnant chylomicron is taken into the liver cell via endocytosis
- the vesicle fuses with lysosomes where the contents are degrades the contents
Early cleavages of the embryo
- the zygote undergoes a series of mitotic divisions in the first three days as it moves through the fallopian tube
- by 48 hours, 8 blastomeres form without any increase in the size of the embryo due to the zona pellucida (at this stage, blastomeres can be removed for PGD)
- on day 3-4, at the 8-16 cell stage, the morula forms; consists of outer cells that will form the trophoblast and inner cell mass cells; this process is called compaction and it is mediated by E-cadherin
Formation of the blastocyst
- on day 4 the morula arrives in the uterus
- it cleaves and develops into an early blastocyst
- it begins to form an blastoceol filled with fluid from the uterus
- it sheds the zona pellucida and expands in size in a process called hatching
- it continues to divide into a late blastocyst, containing hundreds of cells
Implantation of the blastocyst
- On day 7 it attaches ti the endometrail endothelium
- It becomes further imbedded through days 8-14
- during this time there is differentiation: ICM becomes epiblast and hypoblast and this two-layered structure is the bilaminar embryonic disc
- the trophoblasts also differntiates (will make hCG* and the placenta)
- Implantation is complete at the end of the second week
- *hCG is detected during pregnancy tests
The process of gastrulation
- around day 14, the epiblast cells develop into the three germ layers (endoderm, mesoderm, and ectoderm)
- the whole process of gastrulation only takes 2-3 days but is a critical step for normal development
- errors in this step are a major cause of spontaneous abortions
What are the major causes of spontaneous abortions?
- chromosomal abnormalities
- cleavage problems
- progesterone insuffuciency
B cell development
- come from pluripotent stem cells in the bone marrow
- Pre-B Cells have undergone V-D-J segment rearrangement, has a mu constant region polypeptide, and is committed to becoming a B cell
- Immature B cell has the developed a light chain for IgM so it can now express IgM on its surface
- the cell undergoes negative selection to make sure that it does nto react to self antigen
- Mature B cells have both membrane IgM and IgD via alternative splicing of a single heavy chain RNA transcript; they can leave the bone marrow and nigrate to secondary lymphoid organs; they are capable of being activated by antigen
What are the types of immunity in which antibodies are involved?
- Passive (receiving pre-formed antibody):
- 1. Natural-maternal IgG passed to the newborn and IgA in the milk
- 2. Artificial- injection of immunoglobulins (rabies immune globulin, intravenous immunoglobulin, or monoclonal antibody)
- Active (adaptive immune response to antigen):
- 1. Natural- infection with pathogen
- 2. Artificial- immunization with a live or attenuated vaccine
What is IVIG?
Intravenous immunoglobulin: a large collection of purified serum antibodies from very large pools of donors used to treat individuals deficient in humoral immunity
What is Transient Hypogammaglobulinemia?
-the phase in which an infant has not yet developed their own IgGs but is running out of their passively acquired ones from mom; they are at an increased risk for infection
How do you make monoclonal antibodies?
- they are an essentially unlimited supply of antibodies produced by a single clone of B cells
- produced in hybridomas, the result of fusing immortal cancer cells with B cells producing antibody
- -chimeric antibodies: genetically engineered to have mouse variable regions but human constant regions; nomenclature ending is "ximab"
- -humanized antibodies: engineered to have only CDRs from the antigen-specific mouse antibody grafted into a human immunoglobulin sequence; nomenclature ending is "zumab"
- -fully human mAbs: make a library of VH and VL genes; attach these to macrophages and expose them to antigen of interest; grow the ones that bind to the anitgen; nomenclature ending is "umab"
What are the three types of bone cells?
- Osteoblasts: synthesize the organic components of the bone matrix; located exclusively on the surface of the bone matrix
- Osteocytes: individual osteoblasts that become surrounded in their own secretions enclosed within lacunae
- Osteoclasts: monocyte derivitive; multinucelated giant cell that resorbs bone
Classification of lymphoid tissues
WHat are the two types of bone formation?
- 1. Endochondral Ossification: hyaline cartilage model of the bone is laid down first, then filled in with bone tissue
- 2. Intramembranous Ossification: mesenchymal tissue differentiates into osteoblasts which begin the bone formation process
What are the four layers of differntiation at the epiphyseal growth plate?
- From the cartilage towards the bone:
Where are the three different types of cartilage found?
- Hyaline cartilage: respiratory tract and nose
- Elastic cartilage: ear , larynx, epiglottis
- Fibrocartilage: intervertebral discs
WHat are the histological features of different types of arteries?
- Large, elastic (Aorta): Initma is simple squamous, media is all elastic fibers (little/no muscle) with vasa vasorum, and the adventitia is normal
- Medium, muscular artery: intima has a prominent internal elastic membrane, dominantly smooth muscle (more than 5 layers) surrounded by the external elastic membrane, adventitia is thicker
- Arteriole: intima has IEM that is difficult to see, media has 1-5 layers of smooth muscle and no EEM, adventitia blends into surrounding tissue
Where are the three types of capillaries located?
- Continuous: muscle and skin
- Fenestrated: kidney and endocrine tissue
- Discontinuous: liver and spleen
What are the three layers of the heart?
- Endocardium: inner-most layer next to lumen, contains purkinje fibers
- Myocardium: layer of cardiac muscle
- Epicardium: outer-most layer, a sac that holds the heart, aka pericardium
What are the layer sof the epidermis (apical to basal)?
- 1. Stratum Corneum: stratified squamous keratinizing epithelium
- 2. Stratum Lucidum: thin, translucent layer of flattened eosinphilic cells; only seen in thick skin
- 3. Stratum Granulosum: full of granules of lamellar bodies (lipids) that waterproof skin
- 4. Stratum Spinosum: usually the thickest layer; polyhedral cells with desmosomes and tonofibril; actively synthesize keratin
- 5. Stratum Basale: single layer of columnar or cuboidal cells; serve as stem cells for all other layers; many desmosomes and hemidesmosomes; has melanocytes which produce melanin pigment
What is melanin?
- pigment in the skin dervied from tyrosine
- made by melanocyts (neural crest cell derivatives) that look like they have fingers
- it protects keratinocytes from UV damage so it aligns on the apical surface of the cell that faces the sun
- all people have the same number of melanocytes, they are just more active in people with darker skin
What is hair?
- elongated keratinized structures derived from invaginations of the epidermal epithelium called hair follicles
- at the base is the dermal papilla which contains a capillary network required to sustain the hair follicle; this must be destroyed for permanent hair removal
- closely associated with subaceous glands
What are the types of glands found in the skin?
- Eccrine: forehead, armpits, and palms; merocrine secretion (exocytosis in vesicle); for thermoregulation and salt balance; narrow lumen; myoepithelial contractile cells squeeze the sweat out
- Apocrine: mostly in the axillary and perineal regions; wider lumen; secretion by blebbing; also have myoepithelial cells
- Subaceous: discharges sebum (TAGs, FAs, and cholesterol) into the lumen of the hair follicle; holocrine secretion (cell lysis); nuceli only seen on one side of the gland
Histological differences in the respiratory tract
What is a Clara Cell?
- dome-shaped, non-ciliated cells in the bronchioles
- make and secrete surfactant
- controls the chloride transport
- mteabolizes inhaled chemicals
What is the difference between Type I and Type II alveolar cells?
- Type I: line the alveolar surface, simple squamous
- Type II: cuboidal cells in the corners of the alveoli; make surfactant for the alveoli; serve as stem cells if there is damage to the lung; surfactant stores in multilamellar bodies
What are pores of Kohn?
pores between the alveoli that allow venitaltion
Life cycle of post-absorptive lipoproteins
- VLDL's of TAGs and some CE are assembled in a hydrohobic pocket of the ER in hepatocytes; it is associated with apoprotein B100, then acquired Apo C-II and Apo E from HDL once in the blood stream
- Apo C-II allows it to be a substarte for Lipoprotein lipase (LPL) so TAGs are removed and Apo C-ii is lost to form an IDL
- IDL (can be absorbed into liver cells by binding LDL receptors) or it is acted upon by HTGL on the surface of the liver cells, further depleting it of TAGs; ot loses Apo E and it is now an LDL
- LDL is the major serum transporter of cholesterol; has the Apo B100 which is easilt recognized by the LDL receptor on the surface of cells that are thristy for cholesterol
Why is excess LDL a bad thing?
- if it accumulates it can become oxidized and exported out of the lumen of the blood vessel
- once in the intima it recruits leukocytes and stimulates and inflammatory response
- macrophages differentiate into foam cells and begin to die; they release growth factors which causes proliferation of the smooth muscle of the vessel
What is HDL? And why is it the "good cholesterol"?
- high density lipoprotein assembled in the liver with Apoproteins A-I, C-II and E
- transfers Apo C-II and E to other lipoproteins when they are needed
- collects extra cholesterol from cells by converting it to cholesterol esters using the LCAT on the Apo A-I and packages it into its hollow core
- it can also pick up TAGs by exchanging them for CEs with chylomicrons, VLDL, and IDL using CETP
- it then binds to scevenger receptors in th eliver and adrenals so the stuff inside can be process and used
- it is considered "good" because it promotes cholesterol efflux from the vessel walls
What regulates the breakdown of Fatty Acids in the adipose tissue for energy?
- glucagon and epinephrine activate hormone-sensitive lipase (HSL) in adipose tissue which hydrolyzes stored TAGs to release FAs
- FAs are transported through the blood while bound to serum albumin which carries it to other cells to be used to generate energy
Fatty acid catabolism
- Step 1: free FAs are activated in the cytoplasm to generate Acyl-CoA that can be used for beta-oxidation
- Step 2: Acyl-CoA is esterified by carnitine Acyltransferase I (CAT-I) to form acyl-carnitine <-- major regulatory step; CAT-I inhibited by malonyl-CoA (this is only needed for long chain FAs, short/med can pass through the IMM)
- Step 3: acylcarnitine is transported to the mitochondrial matrix via carnitine acylcarnitine translocase
- Step 4: CAT-II regenerates acyl-CoA and carnitine
- Step 5: reduction of the beta carbon to form a trans double bond; FAD becomes FADH2
- Step 6: beta carbon becomes a hydroxyl group
- Step 7: hydroxyl group becomes a ketone and NAD becomes NADH
- Step 8: the first two carbons are cut off as acetyl-CoA which can be sued in the TCA cycle, and the longer chain feeds back into the cycle
Potential probelms with fatty acid catabolism
- 1. energy yield required
- 2. may have unsaturated or branched fatty acids that need ot be converted by differnt enzymes
- 3. may result in propionly CoA which can be toxic to hte brain
Ketone Body Formation
- Step 1: 2 acetyl CoA molecules are fused to form acetoacetyl CoA
- Step 2: HMG CoA synthase turns one ketone into a hydroxyl group to make HMG CoA (and adds another acetyl CoA
- Step 3: an acetyl CoA is lost to form acetoacetate
- Step 4: this becomes acetone via another intermediate and the oxidation of NADH to NAD
- Note: acetoacetate is soluble and can be transported in the blood; the brain needs ketones for energy when it runs out of glucose and fatty acids can't cross the BBB
What is the respiratory quotient (RQ)?
- a measure of how much CO2 is released by a given tissue on catabolic reactions per O2 that is consumed
- low RQ means that a tissue primarily uses FAs and/or proteins for energy
- high RQ means it uses either glucose or ketone bodies
What are the causes of female infertility?
- 1. Ovulation defects: can becaused by hypothalamic abnormalities like changes in GnRH release, reduced secretion of FSH and LH from the AP, stress, extreme athletic activity, eating disorders and thyroid or adrenal hormonal imbalances
- 2. Polycystic Ovary Syndrome (PCOS): increased levels of LH relative to FSH; most common hormonal disturbance in women of child bearing age
- 3. Abnormalities of the femal reproductive tract that interfere with transport of gametes of embryos or blastocyst implantation: endometriosis, PID, tubal obstructions, uterine abnormalities like fibroids, absence of the uterus (MRKH syndrome), excessive scar tissue from surgery (Asherman's Syndrome), or excessive scar tissue from STIs
What are the top causes of male infertility?
- 1. Testicular defects
- 2. Hormonal abnormalities in the HPO axis
- 3. Abnormally high temperature in the testis due to varicocele in the scrotum
- 4. Damage to or congenital loss od the vas deferens
What is the standard order of fertility treatments?
- 1. Orally administered fertility drugs AKA clomiphene citrate to induce ovulation
- 2. Try intrauterine insemination (IUI) with above drug
- 3. 3 or more cycles of injectable fertility drugs such as hMG or FSH
- 4. 3 or more cycles of IVF
What happens when an APC is activated via binding of antigen to Toll Like Receptor?
- 1. IL-1 and TNF are released: stimulate chemokine production and increase adhesive properties of the endothelium so circulating leukocytes binds and exit the circulation; stimulate production of CRP by the liver which acts on the brain to produce fever
- 2. INF alpha and beta: anti-virals
- 3. Make chemokines: brings cells to the site of infection
- 4. IL-12: activates TH1 and NK cells
- 5. IL-10: self-inhibition
- 6. IL-6: activates bone marrow to make more white blood cells
Which HLA regions are associated with each MHC complex?
- Class I: B, C, and A
- Class II: DP, DQ, DR
Which MHC complexes are found where?
- Class I: on every cell of the body except for RBCs
- Class II: only on APCs
What are the structures of the MHC complexes?
- Class I: transmembrane glycoprotein called the alpha chain non-covalently associated with beta-2 microglibulin
- Class II: two transmembrane glycoproteins called the alpha and beta chains
What diseases are known to be associated with a certain HLA?
- 1. Ankylosis spondylitis; B27
- 2. Narcolepsy; DR2
- 3. Rheumatoid Arthritis; DR4
De novo fatty acid biosynthesis
- Step 1: Citrate is pasively transported down its concentration gradient from the mitochodria to the cytoplasm
- Step 2: Citrate lyase breaks it into actely CoA and OAA in the cytoplasm
- Step 3: (committed, regulated step) Acetly CoA carboxylase (needs biotin) makes malonyl CoA
- Step 4: acyl group of an acetyl CoA is bound to phosphopantethein (Pan) on the FAS enzyme
- Step 5: Acyl transferase moves it to the cystein residue (Cys)
- Step 6: malonyl transferase attaches the malonyl group of malonyl CoA to the Pan
- Step 7: the carbon that is double-bonded to the oxygen in the acyl attached to the Cys, condenses with the malonyl on the Cys; CO2 is released from the malonyl and a four carbon beta-ketone is now attached to Pan with the carbons from the acyl being terminal
- Step 8: the beta-ketone is reduced to form a beta-hydroxyl; uses an NADPH and releases NADP
- Step 9: the molecule is dehydrated to form a 2,3 trans double bond
- Step 10: the double bond is reduced to form a single bond; uses NADPH and releases NADP
- Step 11: this chain is then transfered to the Cys by acyl transferase and the cycle continues until it makes palmatate (16 carbons) and it is cleaved by thioesterase; in mammary gladns, we make fatty acids of C8-C14
Regulation of de novo fatty acid biosynthesis
- regulated step is the ofrmation of malonyl CoA by acetyl CoA carboxylase; this enzyme needs biotin, and is only active as a polymer made up of identical subunits
- inactivated via depolymerization by: fatty acyl CoA, excess malonyl CoA, and by phosphorylation via glucagon/epinephrine pathway using ptoein kinase A
- activated via polymerization by: cytoplasmic citrate, and a protein phosphatase
- Note: FAS is also regulated both at the level of substrate availablility and by adaptive changes in the synthesis of the enzyme
What are the two sources of NADPH that is needed for FA synthesis?
- 1. Pentose-phosphate pathway: two molecules of NADPH made
- 2. NADP-linked malate dehydrogenase (malic enzyme): OAA to malate then pyruvate; moves 2 electrons from NADH to NADPH
Fatty Acid elongation
- In the mitochondria: palmatoyl CoA + acetyl CoA--> beta-keto-steraoyl CoA
- On the Smooth ER: palmotoyl CoA + malonyl CoA --> beta-keto-stearoyl CoA + CO2
- the beta-ketones are the reduced, dehydrated, and reduced again to form a saturated acyl CoA; this can be repeated many times to make a long chain
- note: same process as FA biosynthesis, but differnt enzymes; adds carbons at the carbonyl end so the omega number does not change
Fatty acid desaturation
- the enzyme desaturase is found on the SER
- needs NADH, moleculare oxygen and cytochrome b5
- can add double bonds 3 carbons apart but must be farther than 7 carbons from the terminal methyl carbon; that is why we need the essential fatty acids, we cannot synthesize them on our own
What are our two most importnant essential fatty acids that we must get in our diet?
- both are needed in our membranes and as precursors for intracellular signalling molecules
- alpha-linolenic acid: omega-3 fatty acid; 18:3 (delta)9,12,15
- linoleic acid: omega-6 fatty acid; 18:2(delta)9,12
Biosynthesis of TAGs in Liver and Adipose tissue
- Precursors: glycerol-3-phosphate (from dihydroxyacetone phosphate or from glycerol) and fatty acyl CoA
- Process: the 1 and 2 positions of the G3P are acylated with long chain fatty acyl CoA's; dephosphorylation of the 3 position; acylation of the 3 positions
ds DNA virus
- Early synthesis: moves to nucleus, mRNA, then early proteins
- In vitro translation assay: nothing
- Transfection assay: genome is infectious
- Enzymatic assay: nothing
- Enzymes carried with the virus: nothing
- Enveloped: no
- Host cell lysis: yes
- Structure: icosahedral
- Comments: can cause cancer if its genome is integrated and E1A (Rb) and E1B (p53) are expressed; has a terminal protein as a primer; makes a polyprotein then does differential splicing
- Example: adenovirus
plus strand RNA virus (mRNA)
- Early synthesis: viral proteins in the cytoplasm
- In vitro translation assay: yes
- Transfection assay: yes
- Enzymatic assay: nothing
- Enzymes carried with host: nothing
- Enveloped: no
- Host cell lysis: yes
- Structure: icosahedral
- Comments: VpG protein used as primer; encodes a protease, a replicase, the VpG and viral proteins (VPs) needed for capsid assembly
- Example: Picornaviruses/Polio
negative strand RNA virus
- Early synthesis: plus strand mRNA, viral proteins
- In vitro translation assay: nothing
- Transfection assay: nothing
- Enzymatic assay: makes mRNA
- Enzymes carried with virus: transcriptase
- Enveloped: yes
- Host cell lysis: no
- Structure: helical capsid with envelope
- Comments: HA for adsorption, NA degrades receptor, steals CAP from host to serve as primer, nucleoprotein associated with genome
- Example: Myxovirus/influenza
plus strand RNA (not mRNA) viruses
- Early synthesis: ds DNA
- In vitro translation assay: viral proteins
- Transfection assay: nothing
- Enzymatic assay: makes DNA but not mRNA
- Enzymes with virus: reverse transcriptase
- Enveloped: yes
- Host cell lysis: no
- Structure: icosahedral
- Comments: has a tRNA as a primer; reverse transcription makes LTRs; also carries integrase; can cause cancer if integrated near proto-oncogene
- Example: retrovirus, HIV
What are the two classes of membrane lipids?
- 1. Phospholipids: glycerol (glycerophospholipid) or sphingosine (sphingolipid) backbone with a charged phosphate group
- 2. Glycolipids: sphingosine backbone with an O-linked sugar attached
- Note: all glycolipids are sphingolipids, but not all sphingolipids are glycolipds, may be a phospholipid
What is a glycerophospholipid?
- a glycerol backbone with two fatty acids linked in the 1 and 2 position via an ester linkage and a phosphodiester linkage on the 3 position
- can have long chain alcohols attached to the one position via ether linkage
- there may be head group alcohols attached via the phosphate (NOTE: if there is no alcohol attached here it is a phosphatidic acid, PA)
- possible head group alcohols: choline (tertiary amine), serine (carboxyl and amine at end), ethanolamine (primary amine), glycerol (three alcohols), inositol (ring with 6 hydroxyl groups)
What is cardiolipin?
- glycerol backbone with phosphotidyl groups on both the 1 and the 3 carbons; "double phospholipid"
- common in mitochondrial membranes
CDP-DAG pathway of glycerophospholipid synthesis
- Step 1: activation of the phosphatidic acid (PA) by CTP to form CDP-DAG
- Step 2: the haydroxyl from the head group alcohol attacks the bind between the two phosphate groups and displaces CMP
- This process is used to make: PI, PG, and cardiolipin
CDP-alcohol pathway of glycerophospholipid synthesis
- Step 1: the hydroxyl group of the head group alcohol is phosphorylated by a kinase
- Step 2: CTP activates the above formed molecule to yield CDP-(head group)
- Step 3: the hydroxyl group of DAG (formed from PA) displaces CMP to generate the final product
- This process is used to make: PE, PC, and PS
- NOTE: PC<-->PE<-->PS
What is a plasmalogen?
- Glycerophospholipid with fatty alcohol in ether linkage at C1
- Another example: platelet activating factor (PAF)
- functions: alter mambrane permeability, activate platelet secretion, mediate inflammation, resistnace to oxidative damage in the mitochondrial membranes
What are phospholipases?
- enzymes that can hydrolyze portions of the glycerophospholipid molecule
- Phospholipase A: acts on the ester linkage to release the fatty acid chain; A1 acts on C1 and A2 acts on C2; involved in phospholipid remodeling, hemolytic venoms (lysophospholipids), and eicosanoid signaling (under stress, it cleaves PUFAs which can be made into a variety of eicosanoids)
- Phospholipase C: hydrolyzes the C3 bond on the glycerol side of the phosphodiester bond resulting in DAG and the alcohol phosphate; involved in intracellular second messenger signalling (mostly to regulate inracelular calcium and therefore, cell proliferation)
- Phospholipase D: hydrolyzes the C3 bond on the other side of the phosphodiester bond to liberate PA and a free alcohol head group
How do you make prostaglandins and thromboxanes?
- Step 1: PLA2 releases a long PUFA
- Step 2: cyclooxygenase removes two double bonds and addes two molecules of O2
- Step 3: the above made intermediate becomes PGE2, PGF2, TXA2 and PGI2
- NOTE: cyclooxygenase is inhibited by aspirin and therefore reduces fever and inflammation
How are leukotreines made?
- Step 1: PLA2 releases a long PUFA
- Step 2: lipoxygenase adds only one molecule of O2
- Step 3: the above intermediate becomes HPETE, like leukotreines
- NOTE: these are important in immune cell responses like chemotaxis and cytokine release, they also mediate bronchioconstriction and vascualr permeability
What is a sphingolipid and how is it made?
- has a long chain amino alcohol, sphingosine, as the backbone
- can be acylated on the amino group woth a long chain fatty acyl CoA to form ceramide; note that this has an amide bond
- ceramide can be used to make the choline phospholipid sphingolmyelin which is found in the nervous tissues; this is also the only phingolipid that is a phospholipid
What are glycolipids?
- ceraminde (sphingolipid) conatining a hexose oligosaccharide fromed via and activated sugar
- Glucocerebroside: precursor for gangliosides/globoside; they are important for cell-cell recognition, interaction with symbiotic bacteria in the gut, and the basis for ABO blood groups
- Galactocerebroside: can be modified bu sulfation; sulfatides are a common lipid in the white matter of the brain
- accumulation fo these molecules are a feature os LSDs such as Tay-Sachs
What is the structure of the T-cell Receptor (TCR)?
- composed of an alpha and a beta chain, each with a variable and a constant region and connected with disulfide bonds (a minor subset of t cells have gamma-delta chains)
- has only one antigen binding site
- in non-covalent association with CD3 and zeta polypeptides in order to form the TCR Complex
Which T-cells interact with which MHC complexes?
- MHC Class I: on all nucelated cells and interacts with CD8+ cytotoxic t-cells
- MHC Class II: only on APCs; interacts with CD4+ t-cells
What is the role of CD4 and CD8?
- adhesion molecules; bind to their corresponding MHC to enhance the connection to the cell that is showing antigen
- act as signal transduction molecules
What are CD28 and CD40?
- Costimulator molecules required in addition to MHC and TCR to activate T cells
- CD28 on the t-cell binds to B7 on the APC
- CD40 Ligand (CD154) on the t-cell bonds CD40 on APC
T-Cell development in the thymus
- Double negative cells: have a random arrangment of TCR chains but no CD4 or CD8 molecules; cells expressing gamma or delta leave the thymus for the periphery
- Double Positive cells: have a successful arrangment of alpha and beta and express CD3, CD4, and CD8
- Positive selection: developing T cell becomes educated to the MHC molecules expressed by the thymic cortical epithelial cells; ensures that they recognize self-MHC
- Negative selection: the developing t-cell must interact with an MHC present and use its TCR, CD4 and CD8; this selects out cells that bind too tightly to self-MHC
- Single Positive cells: while in the thymic medulla, the expression of one of the CD molecules is down-regulated; they can then enter the periphery and post up in secondary lymphoid tissue
- occurs in the cytoplasm and the ER mostly in the liver
- Step 1: acetyl CoA gets converted to HMG-CoA by HMG CoA Synthase via an intermediate
- Step 2: it is reduced in the ER by HMG CoA reductase to make mevalonic acid <-- regulatory step (inhibited by high levels of free cholesterol and by phosphorylation via glucagon/epinephrine; and inhibited by statins)
- Step 3: decarboxylated and phosphorylated to make a 5-carbon IPP (an isoprene subunit)
- Step 4: condensation of IPP and its isomer DPP to make 10-C geranyl pyrophosphate
- Step 5: another IPP is added to make a 15-C farnesyl pyrophosphate
- Step 6: head to head reductive condensation of two farnesyl-PP molecules to make 30-C squalene
- Step 7: squalene is oxidized by squalene monoxygenase to form an epoxide
- Step 8: the epoxide undergoes cyclization to make lanosterol
- Step 9: a series of demethylations and rearrangements goves cholesterol
What is the major form of cholesterol break-down?
- via metabolism in the liver to bile salts that can be excreted
- cholesterol is oxidized and modified in a series of reactions to make a variety of bile salts
- they are stored in the gall bladder and secreted through the bile duct into the small intestine to facilitat fat absorption
- they are then actively reabsorbed in the small intestine and shuttled back to the gall bladder to be reused in a process called enterohepatic circulation
What are the important steroid hormones to know?
- Progesterone: from the corpus luteum; cheif hormone during pregnancy
- Testosterone and estradiol: from testes and ovaries; responsible for secondary sex characteristics
- Aldosterone: from the adrenal cortex; regulated electrolyte reabsorption in the kidneys
- Cortisol: from the adrenal cortex; regulated metabolism and suppresses immune response and inflammation
- the aldehyde form is retinal which is located in the retina associated with opsins to make rhodopsins
- it is normally 11-cis but when it absorbs light it become all-trans and it excites the nervous sytem
- it is reisomerized into 11-cis
- retinoic acid, another form of vitamin A is an important intercellular signalling molecule involved in growth and differenetiation
- can be formed de novo via the penultimate intermediate in cholesterol synthesis and UV light
- involved in increasing the intestinal absorption of calcium, bone, formation, and the prevention of rickets in children and osteomalacia in adults
- too much can lead to bone degredation and kidney stones
antioxidant that terminate the peroxidative destruction of PUFAs in tissues
- essential cofactor in the synthesis of prothrombin and several other proteins involved in blood clotting
- abundant in nature and also made by the bacteria in the gut
- target for rat poison and coumadin (stop too much clotting)
What is the difference between endogenous and exogenous protein antigens?
- exogenous: taken up and processed by APCs; associate with MHC Class II
- endogenous: come from inside the presenting cell, generally due to infection by the pathogen; associate with MHC Class I
What is IL-2?
-cytokine produced by CD4 TH1 cells that induces CD8 t-cell proliferation
What are the four major subsets of CD4 t-cells? And what do they do?
- TH1: makes INF-gamma and IL-2; activated macrophages, NK cells, CD8 t-cells and stimulates B cells to produce IgG; activate the classical complement pathway
- TH2: IL-4 induces class switch to IgG; IL-5 activates eisoinophils and when associated with TGF-beta induces class switch to IgA; IL-6 makes the bone marrow make more WBCs; IL-13 induces calss switch ot make IgE
- TH17: IL-17 and IL-22 act on the mucosal sites and neutrophils to induce inflammation when there is a bacterial or fungal infection
What signals induce CD4 t-cell differentiation?
- IL-12 induces TH1 differentiation
- IL-4 induces TH2 differentiation