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Background: Polycystic ovary syndrome (PCOS) is the most common endocrine disorder in reproductive-age women and is associated with high prevalence of insulin resistance (IR) and increases in blood pressure (BP). Activation of the Renin-Angiotensin System (RAS) is implicated in the pathophysiology of the elevated BP in PCOS. Metformin is the most common insulin sensitizer agent used to treat IR in PCOS women. Despite metformin’s ability to inhibit RAS overactivation in non-diabetic models, the potential of metformin to mitigate androgen-mediated RAS overactivation remains unknown. Aim and hypothesis: To test the hypothesis that metformin improves the metabolic abnormalities in PCOS via modulation of the intrarenal RAS. Methods: To induce PCOS, four-week-old female Sprague Dawley rats were implanted with dihydrotestosterone (DHT) pellets (7.5 mg) or placebo (PBO) for 90 days (n=6-10/group). During the final four weeks of DHT treatment, rats received metformin (300 mg/kg/day) in standard chow. Fat, lean mass and HOMA-IR, an index of insulin resistance (IR), were assessed. Plasma adiponectin and leptin levels were quantified by ELISA, while protein levels of angiotensin converting enzyme 2 (ACE2), renin, angiotensin type 2 receptor (AT2R), and angiotensinogen were quantified in the renal cortex by Western-blot. Results: DHT increased body weight (BW) (275.00 ± 8.20 vs 241.80 ± 2.95 g, p Supported by NIH-NIGMS grant P20GM121334 (L.L.Y.C. and D.G.R.), NIH-NIDDK grant R21DK113500 (D.G.R.), NIH-NIGMS grant P20GM121334, P20GM104357, NIH-NHLBI grant P01HL51971, and University of Mississippi Medical Center Medical Student Research Program (R.C.) and the Robert M. Hearin Foundation (R.C.). This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Climate warming in the Arctic and the thawing of frozen carbon stocks are leading to uncertainty as to how bacterial communities will respond, including pollutant degrading bacteria. This study investigated the effects of carbon stimulation and temperature on soil microbial community diversity and explosive biodegradation in two sub-Arctic soils. Chitin as a labile carbon source stimulated overall microbial activities as reflected by increases in basal respiration (three to tenfold) and potential nitrification activity (two to fourfold) compared to unamended soil. This stimulation extended to 2,4-dinitroluene- (DNT) and hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX)-degrading microorganisms either directly or via co-metabolic reaction mechanisms. A stimulatory effect of the incubation temperature (2, 12, or 22 °C) on these microbial activities was also observed, but the chitin stimulation caused greater shifts in the structure of the bacterial and fungal communities. The first reported occurrence of an associated role of chitinolytic bacteria belonging to Cellulomonadaceae and chitinolytic fungi belonging to Mortierellaceae in explosive biodegradation is described. This study found that sub-Arctic soil microbial communities were adapted to respond quickly to an increase in labile carbon sources over the range of temperatures used in this study. The warming climate in the Arctic could benefit explosive contaminated soil clean-up by providing non-recalcitrant carbon sources that stimulate overall microbial activity and correspondingly explosive biodegradation.

Polycystic ovary syndrome (PCOS) is the most common endocrine disorder in reproductive-age women. PCOS is characterized by hyperandrogenism and ovulatory dysfunction. Women with PCOS have a high prevalence of obesity, insulin resistance (IR), increased blood pressure (BP), and activation of the renin angiotensin system (RAS). Effective evidence-based therapeutics to ameliorate the cardiometabolic complications in PCOS are lacking. The sodium-glucose cotransporter-2 (SGLT2) inhibitor Empagliflozin (EMPA) reduces BP and hyperglycemia in type 2 diabetes mellitus. We hypothesized that hyperandrogenemia upregulates renal SGLT2 expression and that EMPA ameliorates cardiometabolic complications in a hyperandrogenemic PCOS model. Four-week-old female Sprague Dawley rats were treated with dihydrotestosterone (DHT) for 90 days, and EMPA was co-administered for the last three weeks. DHT upregulated renal SGLT2, SGLT4, and GLUT2, but downregulated SGLT3 mRNA expression. EMPA decreased DHT-mediated increases in fat mass, plasma leptin, and BP, but failed to decrease plasma insulin, HbA1c, or albuminuria. EMPA decreased DHT-mediated increase in renal angiotensin converting enzyme (ACE), angiotensin converting enzyme 2 (ACE2), and angiotensin II type 1 receptor (AGT1R) mRNA and protein expression. In summary, SGLT2 inhibition proved beneficial in adiposity and BP reduction in a hyperandrogenemic PCOS model, however, additional therapies may be needed to improve IR and renal injury.

Natural communities of microbes inhabiting amphibian skin, the skin microbiome, are critical to supporting amphibian health and disease resistance. To enable the pro-active health assessment and management of amphibians on Army installations and beyond, we investigated the effects of acute (96h) munitions exposures to Rana pipiens (leopard frog) tadpoles and the associated skin microbiome, integrated with RNAseq-based transcriptomic responses in the tadpole host. Tadpoles were exposed to the legacy munition 2,4,6-trinitrotoluene (TNT), the new insensitive munition (IM) formulation, IMX-101, and the IM constituents nitroguinidine (NQ) and 1-methyl-3-nitroguanidine (MeNQ). The 96h LC50 values and 95% confidence intervals were 2.6 (2.4, 2.8) for ΣTNT and 68.2 (62.9, 73.9) for IMX-101, respectively. The NQ and MeNQ exposures caused no significant impacts on survival in 96h exposures even at maximum exposure levels of 3560 and 5285 mg/L, respectively. However, NQ and MeNQ, as well as TNT and IMX-101 exposures, all elicited changes in the tadpole skin microbiome profile, as evidenced by significantly increased relative proportions of the Proteobacteria with increasing exposure concentrations, and significantly decreased alpha-diversity in the NQ exposure. The potential for direct effects of munitions exposure on the skin microbiome were observed including increased abundance of munitions-tolerant phylogenetic groups, in addition to possible indirect effects on microbial flora where transcriptional responses suggestive of changes in skin mucus-layer properties, antimicrobial peptide production, and innate immune factors were observed in the tadpole host. Additional insights into the tadpole host's transcriptional response to munitions exposures indicated that TNT and IMX-101 exposures significantly enriched transcriptional expression within type-I and type-II xenobiotic metabolism pathways, where dose-responsive increases in expression were observed. Significant enrichment and increased transcriptional expression of heme and iron binding functions in the TNT exposures served as likely indicators of known mechanisms of TNT toxicity including hemolytic anemia and methemoglobinemia. The significant enrichment and dose-responsive decrease in transcriptional expression of cell cycle pathways in the IMX-101 exposures was consistent with previous observations in fish, while significant enrichment of immune-related function in response to NQ exposure were consistent with potential immune suppression at the highest NQ exposure concentration. Finally, the MeNQ exposures elicited significantly decreased transcriptional expression of keratin 16, type I, a gene likely involved in keratinization processes in amphibian skin. Overall, munitions showed the potential to alter tadpole skin microbiome composition and affect transcriptional profiles in the amphibian host, some suggestive of potential impacts on host health and immune status relevant to disease susceptibility.

Introduction: Polycystic ovary syndrome (PCOS) is the most common endocrine disorder in women of reproductive age. PCOS is characterized by androgen excess and ovulatory dysfunction high prevalence of cardiovascular risk factors such as increased blood pressure (BP), insulin resistance (IR), and obesity. We have demonstrated previously that exposing prepubertal female rats to dihydrotestosterone (DHT) leads to increase in food intake (FI), body weight (BW), BP, and IR. We tested the hypothesis that administration of the AR blocker bicalutamide (BICA) would decrease BP, IR, and obesity in PCOS model. As there are previous reports of severe hepatotoxicity with the AR blocker flutamide, we also examined BICA effects in the liver. Methods: Four-week old female Sprague Dawley rats implanted with DHT pellets (7.5mg/90 days) or placebo (PBO) were randomized to standard chow diet with or without the AR blocker bicalutamide (BICA) at a dose of 250 mg/kg/day throughout the study (n=10/group). BW and FI were measured weekly. BP and heart rate (HR) were measured by radiotelemetry. Fasting plasma was collected for IR (Homeostatic model assessment for IR, HOMA-IR). At euthanasia, the liver was collected, as well as plasma for gamma glutamyl transferase (GGT), alanine transaminase (ALT), and aspartate transaminase (AST) quantification. Results: PCOS rats had increased BW, FI, IR, and BP compared to PBO. BICA treatment had no impact on BW (285.3 ± 7.0 vs 270 ± 8.2 g, P=0.2) as well as FI and HR in PCOS. However, in PCOS, BICA decreased HOMA-IR (5.10 ± 0.40 vs 3.33 ± 0.31, P

Analysis of microbial communities is of broad interest in biology and high throughput sequencing is now the preferred method. However, some studies may not need the level of detail high throughput sequencing provides and its cost may limit the number of samples that can be sequenced. High resolution melting analysis (HRM) of 16S rRNA gene variable regions has been proposed as an efficient and low cost method to prioritize samples for sequencing but more specific primers are needed and its efficacy needs to be confirmed. We tested a more specific pair of primers and compared results concerning the structure of microbial communities in tadpole intestine and feces obtained using HRM, denaturing gradient gel electrophoresis (DGGE) and high-throughput sequencing performed in parallel. All three methods showed similar conclusions concerning the communities and revealed communities that differed among intestinal regions and feces. The improved HRM method targets a shorter amplicon in the V3 region of the 16S rRNA gene and uses non-degenerate primers, both of which increase the sensitivity of HRM. The HRM approach was shown to be as effective as DGGE for comparing microbial communities, is considerably easier to perform, and can be used to assess differences in microbial community structure among a large number of samples before committing to sequencing.

Microbiome studies focused on ecologically relevant vertebrate models like reptiles have been limited. Because of their relatively small home range, fast maturation, and high fecundity, lizards are an excellent reptilian terrestrial indicator species. For this study we used the green anole, Anolis carolinensis, to assess the impact of military relevant contaminants on fecal microbiome composition. Fourteen day sub-acute exposures were conducted via oral gavage with 2,4,6-Trinitrotoluene (TNT) and inorganic lead at doses of 60 mg/kg and 20 mg/kg of body weight, respectively. Body weights and food consumption were monitored and fecal samples were collected for high-throughput 16S rRNA gene amplicon sequencing and analytical chemistry at days 0 and 15. At the end of the study, liver and gut were harvested for body burden data. Chemical analysis confirmed accumulation of TNT, TNT transformation products, and lead in liver tissue and fecal samples. Bacterial community analysis of fecal material revealed significant differences between day 0 and day 15 of TNT exposed anoles with an operational taxonomic unit (OTU) within the genus Erwinia representing 32% of the microbial community in TNT exposed anoles. Predictable changes in gut microbiome composition could offer an easily assayed, noninvasive biomarker for specific chemical exposure providing enhanced scientific support to risk assessments on military installations.