Cytokine Expression After Vaginal Distension of Different Durations in Virgin Sprague Dawley Rats

Abstract

Purpose-To investigate the effect of duration of vaginal distention (VD) on the differential expression of stem cell homing and tissue repair cytokines and cytokine receptors in order to identify factors most important for recovery from injury.Methods-Twenty 10-week-old virgin Sprague-Dawley rats were divided into 4 groups: 1-hour VD, 4-hour VD, 6-hour VD, and anesthetized shams. Vagina, bladder, urethra and rectum were harvested immediately after VD and Real-Time PCR was used to determine relative expression of cytokines and receptors of interest. Mixed models analysis was used to determine associations between expression level and VD duration.Results-Positive associations between VD duration and level of urethral expression were found for one of the receptors of MCP-3 [CCR1 (p=0.0001)], as well as, for MCP-3 (p=0.025), CCR5 (p=0.032), and HIF1-alpha, (p=0.023). A positive relationship between VD duration and MCP-3 expression was also observed in rectal tissue (p=0.035). Urethral expression of CCR2, another receptor for MCP-3, approached significance (p=0.066). An inverse relationship between VD duration and expression of IL8 was found in the bladder (p=0.0008). No association was demonstrated between VD duration and expression of SDF-1, CXCR4, CCR2, CCR3, and VEGF in any pelvic organs.Conclusions-These data support a relationship between VD duration and subsequent expression of MCP-3 and one of its associated receptors (CCR1) in the urethra immediately following VD. The increase in HIF1-alpha expression in the urethra with prolonged VD suggests a limited role of tissue ischemia in the immediate response of pelvic organs to VD.

Simulated childbirth injuries in an inbred rat strain

“…Recent studies have shown that SDF-1 levels are related to the degree of hypoxia, such that hypoxic gradients in tissues directly correlate with SDF-1 gene expression (28). SDF-1 was significantly under-expressed immediately after VD in all tissues in SD animals (17;18); however, it only showed a significant difference in the urethra and vagina, but not rectum and bladder in Lewis rats immediately after VD compared to control rats, suggesting that Lewis rats might have a different pattern of hypoxia after VD, consistent with experiments studying ischemia after stroke (27). Delayed expression of SDF-1 after VD is a possibility and will be investigated in future experiments.…”

“…MCP-3 quantification in Lewis rats following VD demonstrated overexpression patterns in the urethra and vagina similar to that of SD rats, suggesting similar gene expression profiles between the two strains (17;18). …”

“…Also similar to SD rats, Lewis rats were found to have decreased levels of SDF-1 in urethral and vaginal tissues after VD (17;18). SDF-1 is a known homing molecule for hematopoietic stem cells and lymphocytes to various organ systems, including myocardium after myocardial infarction, brain after stroke and synovial tissues in the setting of rheumatoid arthritis (16;27).…”

“…We have previously demonstrated significant upregulation of the stem-cell homing cytokine, MCP-3 in urethral and vaginal tissues following VD in SD rats (17;18). MCP-3 quantification in Lewis rats following VD demonstrated overexpression patterns in the urethra and vagina similar to that of SD rats, suggesting similar gene expression profiles between the two strains (17;18).…”

“…Total RNA was isolated from frozen tissues using RNeasy Mini RNA isolation kit (Qiagen, Valencia, CA) as we have previously described (17;18). RNA concentration was determined using a Ribogreen assay (Molecular Probes, Eugene, OR).…”