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#immune response

50 public questions tagged with this topic.

Vaccine

mRNA vaccines work by:

Messenger RNA vaccines represent nucleic acid platform delivering synthetic mRNA encoding pathogen antigen directly to host cytoplasm where host ribosomal machinery translates it transiently. Manufacturing involves in vitro transcription using T7 RNA polymerase from linearized plasmid DNA template encoding spike protein, incorporation of modified nucleoside N1 methylpseudouridine replacing uridine reducing TLR7 mediated inflammation and increasing translation half-life, addition of 5' cap 7-methylguanosine via vaccinia capping enzyme and 2'O-methylation, and 3' polyadenine tail 100 nt for stability. mRNA encapsulated in lipid nanoparticles LNP composed of ionizable lipid ALC-0315 or SM-102 protonated at low pH encapsulating RNA, helper lipid DSPC forming bilayer, cholesterol fluidity, and PEGylated lipid reducing aggregation and prolonging circulation. Upon intramuscular injection LNPs fuse with cell membrane or endocytosed, ionizable lipid becomes cationic at endosomal low pH disrupting endosome releasing mRNA to cytosol. Ribosomes translate antigen protein undergoing post translational modifications, secreted or membrane anchored, processed for MHC I and II presentation inducing neutralizing IgG and CD4 Th1 and CD8 cytotoxic memory. mRNA degraded within days by RNases no integration risk enabling rapid design.

Ref: Pardi et al. Nat Rev Drug Discov 2018 mRNA mechanism; Polack NEJM mRNA; CDC mRNA work.

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Immunological Applications

Subunit vaccines contain:

Whole pathogen vaccines contain complex mixture of antigens including immunodominant protective components but also many irrelevant proteins plus reactogenic factors such as lipid A endotoxin activating TLR4 causing fever, pertussis toxin ADP-ribosylating Gi, increasing reactogenicity and rare adverse events. Subunit vaccine strategy improves safety by purifying protective antigens through biochemical fractionation, recombinant expression, or synthetic peptide synthesis excluding toxic moieties. Hepatitis B vaccine exemplifies recombinant subunit: hepatitis B small surface antigen HBsAg gene cloned into Saccharomyces cerevisiae yeast downstream of alcohol dehydrogenase promoter, protein expressed self-assembles into 22 nanometer virus-like particles composed of 100 monomers with disulfide crosslinks displaying conformational neutralizing a determinant in second hydrophilic loop, particles lack viral DNA completely noninfectious. Acellular pertussis vaccine includes genetically detoxified pertussis toxin PT9K/129G where arginine 9 to lysine and glutamic acid 129 to glycine mutations abolish ADP-ribosyltransferase activity, filamentous hemagglutinin adhesion factor, and pertactin 69 kDa outer membrane protein, eliminating whole cell lipopolysaccharide linked to high fever and protracted crying observed with whole cell DTP.

Ref: NCERT Class XII Biology subunit purified antigen VLP HBsAg; Pulendran Nat Immunol 2011 alum NLRP3 adjuvant acellular pertussis.

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Gene Therapy

Which viral vector shows very low immune response?

Recombinant AAV produced by triple transfection HEK293 cells with plasmids encoding rep/cap removed from vector, adenoviral helper functions E2A E4 VA RNA, and vector genome flanked by ITRs yields particles depleted viral coding sequences that normally generate PAMPs triggering innate and adaptive immunity. Capsid proteins VP1 VP2 VP3 ratio 1:1:10 from serotypes AAV2 AAV5 AAV9 exhibit low TLR2 mediated NF-kB activation and minimal TLR9 stimulation due to CpG depletion via codon optimization resulting reduced type I interferons, TNF-alpha, IL-12. Consequently transduced hepatocytes, muscle fibers, dorsal root ganglia display prolonged persistence without CTL elimination although capsid-specific CD8 cells can occur at high doses. Humoral neutralizing antibodies predominantly preexisting limits redosing but allows primary treatment. Episomal concatemers persist as circular monomers multimers associated with histones providing transcriptionally active chromatin for years minimal genotoxicity due low random integration

Ref: PMC AAV Low Immunogenicity Review PMCID 10142300; UQ Biomedical Sciences Vector Immunogenicity Comparison; Nature Med AAV Clinical Safety.

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Cell Organelles

Which lysosomal membrane protein is required for fusion with phagosomes?

Phagosome maturation in macrophages and neutrophils converts a nascent bacteria-containing vacuole into a microbicidal phagolysosome through sequential interactions with early endosomes, late endosomes and terminal lysosomes. Acquisition of lysosomal membrane proteins marks progression, with lysosome-associated membrane proteins 1 and 2, LAMP1 and LAMP2, being the most abundant heavily glycosylated proteins delivered to maturing phagosomes, protecting membrane from hydrolases. Studies using LAMP-1 and LAMP-2 knockout mice and combined siRNA show single loss causes mild delay while double deficiency impairs recruitment of Rab7 GTPase, loss of early marker EEA1 and failure of lysosomes to fuse, indicating cooperative requirement for tethering and SNARE complex assembly alongside syntaxin 7 and VAMP8. Additionally, phagosome resolution into lysosome-derived vesicles depends on clathrin coats, actin polymerization via Arp2/3 and microtubule motors that fragment the vacuole after cargo degradation, recycling membrane for reuse. Clathrin adaptors AP1 and EpsinR also deliver lysosomal phosphatases via Rab5 early endosomes to expand phagocytic cup. Thus both structural LAMP proteins and clathrin-mediated membrane remodeling cooperate sequentially in phagolysosome formation and turnover for host defense.

Ref: Huynh et al., EMBO Journal 2007: LAMP Proteins Are Required for Fusion of Lysosomes with Phagosomes.

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