A Systems Biology Examination of the Human Female Genital Tract Shows Compartmentalization of Immune Factor Expression

Abstract

f Many mucosal factors in the female genital tract (FGT) have been associated with HIV susceptibility, but little is known about their anatomical distribution in the FGT compartments. This study comprehensively characterized global immune factor expression in different tissue sites of the lower and upper FGT by using a systems biology approach. Tissue sections from the ectocervix, endocervix, and endometrium from seven women who underwent hysterectomy were analyzed by a combination of quantitative mass spectrometry and immunohistochemical staining. Of the >1,000 proteins identified, 281 were found to be differentially abundant in different tissue sites. Hierarchical clustering identified four major functional pathways distinguishing compartments, including innate immune pathways (acute-phase response, LXR/RXR) and development (RhoA signaling, gluconeogenesis), which were enriched in the ectocervix/endocervix and endometrium, respectively. Immune factors important for HIV susceptibility, including antiproteases, immunoglobulins, complement components, and antimicrobial factors, were most abundant in the ectocervix/endocervix, while the endometrium had a greater abundance of certain factors that promote HIV replication. Immune factor abundance is heterogeneous throughout the FGT and shows unique immune microenvironments for HIV based on the exposure site. This may have important implications for early events in HIV transmission and site-specific susceptibility to HIV in the FGT.

Increased levels of inflammatory cytokines in the female reproductive tract are associated with altered expression of proteases, mucosal barrier proteins, and an influx of HIV-susceptible target cells

“…Reversed-phase liquid chromatography (high pH RP, Agilent 1200 series microflow pump, Water XBridge column) was used for desalting and detergent removal of peptides using a step-function gradient as described previously. 21 Eluted fractions were dried via vacuum centrifugation and kept at À 80 1C until analyzed by mass spectrometry.…”

“…Recent technological advances in mass spectrometry offer the promising opportunity to discover new mechanisms linking inflammatory cytokines and HIV risk in human cohorts, via the concomitant measurement of hundreds of proteins from small volume human mucosal samples. [21][22][23][24] However, collection of data on this scale can lead to complexities of interpretation, as deciphering long ''significant'' protein lists or univariate analyses can miss important effects on pathways and perturbations in function categories. This can be mitigated in part by clustering and pathway analyses tools, which offer the opportunity to understand study group distinctions based on groups of biomarkers.…”

Molecular Signatures of Immune Activation and Epithelial Barrier Remodeling Are Enhanced during the Luteal Phase of the Menstrual Cycle: Implications for HIV Susceptibility

“…Supernatants free of cellular debris were used for all downstream processing. Portions (100 g) of CVL protein, as determined by standard BCA protein assay (Novagen), from each participant were processed according to the protocol used by Birse et al (27,32). In brief, equal amounts of protein from each individual were digested with trypsin and analyzed using a label-free method.…”

“…Indeed, mucosal secretions are exceptionally complex, containing many hundreds of unique factors important for both innate immune defense, as well as inflammatory and anti-inflammatory processes (23,27). It therefore follows that factors that modulate risk of HIV infection are likely complex, involving a network of multiple interacting components.…”

Cataloguing of Potential HIV Susceptibility Factors during the Menstrual Cycle of Pig-Tailed Macaques by Using a Systems Biology Approach

“…These significant tissue architecture modifications might result in gene expression changes which have not been documented for pigtailed macaques to date. Using proteomic analyses, it was shown that there exists an abundance of soluble proteins in different compartments of the human female genital tract (12). Thus, our study goal was to not only catalogue menstrual phase-related changes but also help compare and contrast the pigtail model to women whose fluctuations in susceptibility to HIV during the cycle are unknown.…”