CON Rats Research Articles

Effects of moderate-intensity aerobic training on cardiac structure and function in type 2 mellitus diabetic rats: Based on echocardiography and speckle tracking

PurposeThe present study aimed to evaluate the cardiac function changes using Layer-specific Speckle-tracking echocardiography (LS-STE) induced by Moderate-intensity aerobic training (MIAT) in Type 2 diabetes mellitus (T2DM) rats. MethodsTwenty-six rats were divided into four groups: the Control group (Con), the Training control group (CT), the T2DM group (DM), and the T2DM training group (DT). The CT and DT groups underwent an 8 weeks MIAT. Cardiac structure and function were evaluated by echocardiography and LS-STE. ResultsCompared with the Con group, left atrial diameter (LAD), left ventricular end-diastolic diameter(LVEDD), relation wall thickness (RWT), and left ventricular mass (LVM) were significantly higher, and the endo-, mid-, and epi-longitudinal strain (LS), global radial strain (GRS), and endo- and mid-circumferential strain (CS) were significantly lower in DM rats (all p < 0.05). The endo-, mid-, and epi-LS, GRS, endo- and mid-CS were increased in DT rats compared with DM rats (all p < 0.05). Compared with Con rats, CT rats had a significant increase in LVEDD and LVM (all p < 0.05), meanwhile myocardial strains had no significant differences (all p > 0.05). ConclusionLS-STE was a sensitive method to assess subclinical myocardial changes in T2DM rats. MIAT had the benefit of reversing cardiac systolic subclinical dysfunction in T2DM rats.

Abstract 39: Tryptophan contributes to the sex-dependent loss of diurnal sodium excretion in BMAL1 knockout rats.

Kidney function is time-of-day-(TOD) dependent irrespective of light, sleep, activity, and meal timing. Recent discoveries reveal that diurnal (day/night) kidney function is maintained by an intrinsic molecular clock. We recently observed that loss of this transcriptional regulatory system impairs functional rhythms in electrolyte excretion in a novel rat model. When the core clock gene, brain and muscle ARNT-like protein 1 (BMAL1) is knocked out of Sprague-Dawley rats we now find that male, but not female, BMAL1 knockout (KO) rats no longer have diurnal changes in both glomerular filtration rate (GFR; Day:1.3±0.1 vs Night:1.2±0.1mL/min/100gBW;t-test p=0.919) along with sodium excretion (UNaV) (Day:819±58 vs Night:756±113 μmol/12h; t-test p=0.439) compared to littermate wildtype controls (CON; GFR Day:1.1±0.1 vs Night:1.6±0.1mL/min/100g; t-test p=0.044; UNaV Day: 849±138 vs Night:1423±102 μmol/12h; t-test p=0.003). LC/MS analysis of plasma from KO rats found that levels of tryptophan (trp) were significantly (2-way ANOVA p Interaction =0.847, p&lt;span style="font-size:10.8333px"&gt;Genotype&lt;/span&gt;=0.041, p TimeofDay =0.952) lower in KO males during the day (KO:77±6, CON:99±7μM; Sidak post-hoc p=0.049) but not night (KO:78±5, CON:99±12μM; Sidak post-hoc p=0.084) with no differences in plasma trp seen in KO females (Day:110±26,107±14μM vs Night:96±8,96±12μM KO and CON respectively). Trp had recently been shown to both regulate and be regulated by the circadian clock as well as increase GFR similar to other aromatic amino acids. This led us to hypothesize that TOD trp availability may underlie the changes seen in both UNaV and GFR in male KO rats. Therefore, we gave a single dose of trp (100 mg/kg,PO) at either 7pm (lights off, ZT12) or 7am (lights on, ZT0). After a 1 wk washout period, rats were dosed again in a crossover design. Male KO rats receiving trp at ZT0 had no significant change from baseline in UNaV (818±58 vs 784±61 μmol/12h) over the following 12hrs; however, a significant increase in UNaV (756±114 vs 1152±146 μmol/12h) following ZT12 dosing was observed (2-Way ANOVA p Interaction =0.038, p TrpTime =0.087, p TimeofDay =0.210; Sidak Post-Hoc: baseline vs ZT12 trp p=0.003) with no significant change seen in female KO or either male or female CON rats. Taken together these results suggest that BMAL1 is necessary for maintaining kidney physiologic rhythms in sodium excretion potentially through changes in trp availability.

Butyrate promotes visceral hypersensitivity in IBS model via mast cell-derived DRG neuron lincRNA-01028-PKC-TRPV1 pathway.

Emerging evidence indicates that gut dysbiosis is involved in the pathogenesis of visceral hypersensitivity (VH). However, how gut microbiota contributes to the development of VH is unknown. Here, we sought to examine the signal transduction pathways from gut to dorsal root ganglion (DRG) responsible for this. Therefore, abdominal withdrawal reflex (AWR) scores, fecal output, fecal water content, and total gastrointestinal transit time (TGITT) were assessed in Con rats, VH rats, rats treated with NaB, and VH rats treated with VSL#3. Fecal microbiota and its metabolite (short-chain fatty acids, SCFAs), mast cell degranulation in colon, lincRNA-01028, miR-143, and protease kinase C (PKC) and TRPV1 expression in DRGs were further detected. VH rats showed an increased fecal water content, a shortened TGITT, an increased abundance of Clostridium sensu stricto 1 and increased butyrate in fecal samples, an increased mast cell degranulation, an increased expression of lincRNA-01028, PKC, and TRPV1, and a decreased expression of miR-143 in DRGs compared with control rats, which could be restored by the application of probiotic VSL#3. The above-mentioned detection in rats treated with butyrate was similar to that of VH rats. We further confirm whether butyrate sensitized DRG neurons by a lincRNA-01028, miR-143, and PKC-dependent mechanism via mast cell in vitro. In co-cultures, MCs treated with butyrate elicited a higher TRPV1 current, a higher expression of lincRNA-01028, PKC, and a lower expression of miR-143 in DRG neurons, which could be inhibited by a lincRNA-01028 inhibitor. These findings indicate that butyrate promotes visceral hypersensitivity via mast cell-derived DRG neuron lincRNA-01028-PKC-TRPV1 pathway.IMPORTANCEIrritable bowel syndrome (IBS), characterized by visceral hypersensitivity, is a common gastrointestinal dysfunction syndrome. Although the gut microbiota plays a role in the pathogenesis and treatment of irritable bowel syndrome (IBS), the possible underlying mechanisms are unclear. Therefore, it is of critical importance to determine the signal transduction pathways from gut to DRG responsible for this in vitro and in vivo assay. This study demonstrated that butyrate sensitized TRPV1 in DRG neurons via mast cells in vivo and in vitro by a lincRNA-01028, miR-143, and PKC-dependent mechanism. VH rats similarly showed an increased abundance of Clostridium sensu stricto 1, an increased fecal butyrate, an increased mast cell degranulation, and increased expression of TRPV1 compared with control rats, which could be restored by the application of VSL#3. In conclusion, butyrate produced by the altered intestinal microbiota is associated with increased VH.

Investigation on Transient Receptor Potential Vanilloid 1 (TRPv1) Activation in Skeletal Muscle: Pilot Studies

Transient Receptor Potential Vanilloid 1 (TRPv1) is a Ca2+ ion channel predominately studied in neuronal cells and has been found to have sex-differential expression in mouse arterial smooth muscle. More recently, TRPv1 expression has also been observed in skeletal muscle. Among other stimuli, osmotic stress has shown to activate TRPv1 in different tissues (e.g., retinal ganglion and supraoptic nuclear cells). However, the effect of hyperglycemia- and dehydration-induced osmotic stress and the sex-differential expression on TRPv1 in skeletal muscle is not known. PURPOSES: To examine the effect of sex ( Pilot 1), hyperglycemia ( Pilot 2), and dehydration ( Pilot 3) on TRPv1 in skeletal muscle. HYPOTHESES: We hypothesized that TRPv1 would be different between sexes ( Pilot 1), would be higher in hyperglycemic rats compared to healthy controls ( Pilot 2), and would be greater in dehydrated than hydrated state in men ( Pilot 3). METHODS: To accomplish these purposes, we analyzed a subset of samples from three previously completed projects. Pilot 1: Vastus lateralis (VL) muscles were collected from untrained men ( n=3) and women ( n=3) at baseline. Pilot 2: Rats fed a high fat diet and injected with streptozotocin (35 mg/kg) (HFD; n=3; non-fasting blood glucose (NFBG) ≥300mg/dL) were compared to rats fed a normal diet (CON; n=3; NFBG &lt;300mg/dL). Soleus (Sol) muscle was collected after 7 weeks of feeding. Pilot 3: Lastly, VL muscles were collected from trained men (n=5) either dehydrated (DEHY; 296 ± 6 plasma mmol/kg) or hydrated (EUHY; 283 ± 5 plasma mmol/kg) for 24 hours. Rat Sol and human VL samples were analyzed for TRPv1 and p-TRPv1ser502 protein content. TRPv1 activation was calculated as p-TRPv1ser502/ TRPv1/ total protein. Student’s T-tests and effect sizes (ES; small [≤0.49], medium [0.5-0.79], or large [≥0.8] effect) were used to detect differences. Results: Pilot 1: A significant ( p&lt;0.05) difference was observed for TRPv1 with greater expression in men than women (43%, p=0.03, d=2.757). A medium ES was observed for TRPv1 activation (31%, p=0.42, d=0.728) being greater in men compared to women. Pilot 2: No significant difference was observed for TRPv1 and TRPv1 activation between CON and HFD rats. A medium ES was observed for TRPv1 expression (-9%, p=0.31, d=0.441) being lower in HFD compared to CON. A large effect size was observed for TRPv1 activation (30%, p=0.18, d=1.689) being greater in HFD compared to CON. Pilot 3: No significant difference was observed for TRPv1 and TRPv1 activation between EUHY and DEHY. A large ES was observed for TRPv1 (34%, p=0.15, d=0.995) and small ES was observed for TRPv1 activation (2%, p=0.80, d=0.164) being greater in DEHY compared to EUHY. CONCLUSION: These data are the first to illustrate the potential difference in TRPv1 in skeletal muscle. Future research is needed to explore the impact of differential TPRv1 activation on downstream Ca2+ signaling between sexes, dehydration, and hyperglycemic models. This is the full abstract presented at the American Physiology Summit 2024 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.

Tumor Oxygenation in an Orthotopic Model of Breast Cancer: Impact of Dietary Nitrate Supplementation

Introduction: Breast cancer (BC), the predominant non-melanoma cancer among women, is projected to escalate to more than 3 million new cases and ~1 million fatalities globally by 2040. Chronic and fluctuating low oxygen (O2) conditions within tumors favor malignant growth, decreased responsiveness to therapeutic interventions, and reduced patient survivability. Treatment strategies to combat tumor hypoxia are desperately needed. Dietary nitrate supplementation via beetroot juice (BRJ), which increases nitric oxide (NO) production through the nitrate-nitrite-NO pathway, has been demonstrated to increase skeletal muscle blood flow, improve mitochondrial effciency, and reduce metabolic demands. Specifically, BRJ improves interstitial oxygen pressures (PO2 is) in acidic, low oxygen environments such as that found in the tumor microenvironment. Therefore, we hypothesized that BRJ supplementation would increase PO2 is in BC tumors. Methods: Female Fischer 344 retired breeder rats (6-8 months old) were used in this investigation. Adenocarcinoma cells (MAT B III at 6 x 103) were injected into the mammary duct and tumor growth was monitored over 2 weeks. Once tumors reached ~5 mm in diameter, rats were randomly assigned to consume BRJ (1 mmol/kg/day; BRJ, n=3) or water (CON; n=3) for 5 days. Under pentobarbital anesthesia, tumor PO2 is measurements were obtained using phosphorescence quenching. Results: Tumor growth rate was decreased in BRJ compared to CON rats (0.5 ± 0.1 vs. 1.0 ± 0.1 mm/day; p&lt;0.05). Heart rate and mean arterial pressure were not different between groups (P&gt;0.05). Blood lactate was significantly decreased in BRJ compared to CON rats (0.5 ± 0.1 vs. 1.2 ± 0.1 mmol/L, p&lt;0.05). There was an increase in mean tumor PO2 is in BRJ vs. control rats (6.9 ± 1.3 vs. 3.7 ± 0.47; p&lt;0.05). Conclusions: This study offers promising preliminary insights into the potential of dietary nitrate supplementation as an adjunctive approach to BC treatment to increase tumor PO2 is and reduce tumor growth rate. Future and ongoing studies will increase the sample size, use fluorescent microspheres to measure tumor blood flow, and explore the potential for BRJ and other NO agonists to combat tumor hypoxia and the skeletal muscle dysfunction associated with cancer progression. This research was funded by the following sources: the Johnson Cancer Research Center Innovative Research Award and Sustained Momentum for Investigators with Laboratories Established (SMILE) Grants at Kansas State University, the National Institute on Aging (Grant 1R15AG078060), and Ruth L. Kirschstein National Research Service Awards from the National Heart, Lung, and Blood Institute (Grants 1F31HL167618-01 and 1F31HL170643-01). This is the full abstract presented at the American Physiology Summit 2024 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.

FRI151 Combined Maternal Hyperandrogenemia And Maternal Western Diet Consumption Programs Hypertension In Adult Female Offspring That Is Associated With Increased Leptin Levels

Abstract Disclosure: N.M. Shawky: None. R.M. Quin: None. K. Davenport: None. Funding: P20GM121334, AHA Career Development Award 938320 Background: Polycystic ovary syndrome (PCOS) is the major cause of hyperandrogenemia and the most common endocrinopathy in women of reproductive age. There is still a great controversy on whether daughters of PCOS women develop PCOS themselves. PCOS is commonly associated with increased blood pressure (BP) and obesity. We utilize a hyperandrogenemic female (HAF) rat model that develops a 3-fold increase in androgen levels to study the cardiometabolic consequences in the offspring (off.). We have previously shown that adult female off. of HAF rats consuming regular chow do not develop hypertension (HTN) or obesity. Consumption of western diet (WD) aggravates the reproductive phenotype of PCOS. We have shown that WD consumption also promotes further increase in BP, early onset obesity and hyperleptinemia in HAF rats, an effect that was not seen in normoandrogenemic controls. The current study was performed to test the hypothesis that WD consumption in HAFs (model of PCOS with worsened cardiometabolic phenotype) will program HTN, hyperandrogenemia and obesity in adult female off. Methods: At 4 wks of age, female Sprague Dawley (SD) rats were implanted with either 5α-dihydrotestosterone (7.5 mg, s.c.; HAF) or placebo (controls; CON) pellets, that were replaced every 85 days. HAF and CON rats were alsostarted on WD (high fat and high sucrose) or control diet (CD), thus forming 4 groups; CON-CD, CON-WD, HAF-CD and HAF-WD. At 10-13 wks of age, females were paired with SD males and allowed to get pregnant and lactate their off. At 16 wks of age, body composition (EchoMRI), plasma leptin (ELISA) and serum testosterone (T) [LC/MS] were determined in female off. (n=4-12). Another set of female off. were implanted with radiotelemetry transmitters to measure mean arterial pressure (MAP) [n=4-5]. All studies were done using 1 off./litter/group. Results: Despite the similar fat mass in all groups, plasma leptin was significantly higher in female off. from HAF-WD compared to female off. from CON-WD (1.5-fold, p&amp;lt;0.05). Serum T was significantly higher (p&amp;lt;0.05) in female off. from HAF-CD compared to female off. from CON-CD (4-fold), CON-WD (3-fold) and HAF-WD (3-fold). MAP was significantly higher in female off. from HAF-WD compared to female off. from CON-CD and HAF-CD (113 ± 3 vs 104 ± 2 and 103 ± 5 mmHg, p&amp;lt;0.05). Yet, MAP was similar between female off. from CON-WD, CON-CD and HAF-WD. Conclusion: Our results indicate that independent mechanisms program hyperandrogenemia and HTN in adult female off. Combined maternal hyperandrogenemia and WD could program HTN in female off. that is associated with hyperleptinemia. This suggests that daughters of PCOS women with worsened cardiometabolic phenotype could be at risk of hyperleptinemia and HTN as adults. Our future studies will aim at addressing the role of hyperleptinemia in mediating HTN in the female off. Presentation: Friday, June 16, 2023

Chronic intermittent hypobaric hypoxia improves iron metabolism disorders via the IL-6/JAK2/STAT3 and Epo/STAT5/ERFE signaling pathways in metabolic syndrome rats

AimOur previous study demonstrated that chronic intermittent hypobaric hypoxia (CIHH) improved iron metabolism disorder in obese rats through the downregulation of hepcidin. This study aimed to observe the molecular mechanism of CIHH in improving iron metabolism disorders, especially by Janus kinase/signal transducer and activation of the transcription (JAK/STAT) signaling pathway in metabolic syndrome (MS) rats. MethodsSix-week-old male Sprague-Dawley rats were randomly divided into four groups: CON, CIHH (subjected to hypobaric hypoxia simulating 5000-m altitude for 28 days, 6 h daily), MS (induced by high fat diet and fructose water), and MS+CIHH. The serum levels of glucose, lipid metabolism, iron metabolism, interleukin-6 (IL-6), erythropoietin (Epo) and hepcidin were measured. The protein expressions of JAK2, STAT3, STAT5, bone morphogenetic protein 6 (BMP6), small mothers against decapentaplegic 1 (SMAD1) and hepcidin were examined. The mRNA expressions of erythroferrone (ERFE) and hepcidin were analyzed. ResultsThe MS rats displayed obesity, hyperglycemia, hyperlipidemia, iron metabolism disorder, increased IL-6 and hepcidin serum levels, upregulation of JAK2/STAT3 signaling pathway, decreased Epo serum levels, downregulation of STAT5/ERFE signaling pathway in spleen, upregulation of BMP/SMAD signaling pathway in liver, and increased hepcidin mRNA and protein expression compared to CON rats. All the aforementioned abnormalities in MS rats were ameliorated in MS + CIHH rats. ConclusionsCIHH improved iron metabolism disorders, possibly by inhibiting IL-6/JAK2/STAT3 and activating Epo/STAT5/ERFE signaling pathway, thus downregulating hepcidin in MS rats.

Rats with Long-Term Cholestasis Have a Decreased Cytosolic but Maintained Mitochondrial Hepatic CoA Pool.

Previous studies showed that rats with long-term bile duct ligation have reduced coenzyme A stores per g of liver but maintained mitochondrial CoA stores. Based on these observations, we determined the CoA pool in the liver homogenate, liver mitochondria, and liver cytosol of rats with bile duct ligation for 4 weeks (BDL rats, n = 9) and sham-operated control rats (CON rats, n = 5). In addition, we tested the cytosolic and mitochondrial CoA pools by assessing the metabolism of sulfamethoxazole and benzoate in vivo and of palmitate in vitro. The hepatic total CoA content was lower in BDL than CON rats (mean ± SEM; 128 ± 5 vs. 210 ± 9 nmol/g), affecting all subfractions equally (free CoA (CoASH), short- and long-chain acyl-CoA). In BDL rats, the hepatic mitochondrial CoA pool was maintained, and the cytosolic pool was reduced (23.0 ± 0.9 vs. 84.6 ± 3.7 nmol/g liver; CoA subfractions were affected equally). The urinary excretion of hippurate after i.p. benzoate administration (measuring mitochondrial benzoate activation) was reduced in BDL rats (23.0 ± 0.9 vs. 48.6 ± 3.7% of dose/24 h), whereas the urinary elimination of N-acetylsulfamethoxazole after i.p. sulfamethoxazole administration (measuring the cytosolic acetyl-CoA pool) was maintained (36.6 ± 3.0 vs. 35.1 ± 2.5% of dose/24 h BDL vs. CON rats). Palmitate activation was impaired in the liver homogenate of BDL rats but the cytosolic CoASH concentration was not limiting. In conclusion, BDL rats have reduced hepatocellular cytosolic CoA stores, but this reduction does not limit sulfamethoxazole N-acetylation or palmitate activation. The hepatocellular mitochondrial CoA pool is maintained in BDL rats. Impaired hippurate formation in BDL rats is explained best by mitochondrial dysfunction.

Abstract P048: Western Diet Consumption Exacerbates Hypertension In Hyperandrogenemic Female Rat, A Model Of Polycystic Ovary Syndrome.

Background: Polycystic ovary syndrome ( PCOS ) is the most common endocrinopathy affecting women of reproductive age. It is characterized by hyperandrogenemia, increased blood pressure ( BP ) and metabolic dysfunction. Consumption of western diet ( WD ) aggravates the reproductive and metabolic phenotypes of PCOS. Yet, the effect of WD consumption on BP in PCOS women is not clear. Therefore, this study was undertaken to test the hypothesis that WD consumption in hyperandrogenemic female ( HAF ) rats (a model that mimics most of PCOS symptoms) increases BP along with increasing renal expression of the vasoconstrictor ( VC ) components of the renin-angiotensin system ( RAS ). Methods: At 4 wks of age, female Sprague Dawley rats were implanted (s.c.) with either 5α-dihydrotestosterone pellets (7.5 mg/90 d; HAF rats) or placebo pellets (controls; CON) , that were replaced throughout life. At 4 wks of age, HAF and CON rats were also started on WD (high fat and high sucrose content) or control diet ( CD ), thus forming 4 groups; CON-CD, CON-WD, HAF-CD and HAF-WD. At 20 wks of age, rats were implanted with radiotelemetry transmitters in the abdominal aorta to measure mean arterial pressure ( MAP ) [n=4-5/group]. Another set of rats (n=3-6/group) were euthanized and kidneys collected for measurement of components of the RAS by western blot. Results: are shown in Table 1 . Conclusion: WD consumption in HAF results in further increases in BP, despite upregulation of components of the intrarenal vasodilator RAS (ACE2 and AT2R), which is likely a protective mechanism. Future studies will determine the role(s) of other major VC pathways (e.g. endothelin system) in hypertension in WD-fed HAFs.

Restriction of dietary protein in rats increases progressive-ratio motivation for protein

Low-protein diets can impact food intake and appetite, but it is not known if motivation for food is changed. In the present study, we used an operant behavioral task – the progressive ratio test – to assess whether motivation for different foods was affected when rats were maintained on a protein-restricted diet (REST, 5% protein diet) compared to non-restricted control rats (CON, 18% protein). Rats were tested either with nutritionally-balanced pellets (18.7% protein, Experiment 1) or protein-rich pellets (35% protein, Experiment 2) as reinforcers. Protein restriction increased breakpoint for protein-rich pellets, relative to CON rats, whereas no difference in breakpoint for nutritionally-balanced pellets was observed between groups. When given free access to either nutritionally-balanced pellets or protein-rich pellets, REST and CON rats did not differ in their intake. We also tested whether a previous history of protein restriction might affect present motivation for different types of food by assessing breakpoint of previously REST animals that were subsequently put on standard maintenance chow (protein-repleted rats, REPL, Experiment 2). REPL rats did not show increased breakpoint, relative to their initial encounter with protein-rich pellets while they were protein-restricted. This study demonstrates that restriction of dietary protein induces a selective increased motivation for protein-rich food, a behavior that disappears once rats are not in need of protein.

Ossified Particles in the Peripheral Circulation of Young and Old male Fischer‐344 Rats following Bone Marrow Ablation and Interleukin‐1 Receptor Antagonism

BACKGROUNDA pro‐inflammatory marrow microenvironment presumably augments ossification in bone marrow blood vessels. We speculate that ossifying blood vessels results in bone‐like particles (i.e., ossified particles, OSP) in the peripheral blood. By performing bone marrow ablation (BMA) and administering an interleukin‐1 receptor antagonist (IL‐1RA) during recovery, we sought to reduce the number of OSP in the circulation.METHODSYoung (6‐mon) and old (24‐mon) male Fischer‐344 rats were assigned accordingly: Control (young CON, n=10; old CON, n=8), IL‐1RA (young, n=11; old, n=8), BMA (young, n=12; old, n=7) and combined treatment (young IL‐1RA+BMA, n=8; old IL‐1RA+BMA, n= 8). In the BMA and IL‐1RA+BMA rats, bone marrow in the right femoral shaft was ablated. During 3 weeks of recovery, CON and BMA rats received PBS (100 μL, 3 d/wk, i.p.), while IL‐1RA and IL‐1RA+BMA rats received the antagonist (3 μg/kg, 3 d/wk, i.p.). At sacrifice, left ventricular whole blood samples were collected. Plasma IL‐1α and IL‐1β were measured by ELISA. Five‐hundred microliters of blood were prepared for flow cytometry and OSP were sorted to obtain a count. The diameters of OSP were determined with a Cellometer. Some samples were pooled to meet the minimum detection threshold. These data are expressed as a percent of the total OSP count according to diameter, i.e., % OSP per 0‐14µm, 15‐30µm, 31‐45µm, and 46‐60µm. ELISA data were analyzed by two‐way ANOVA using SPSS (v. 25). The OSP count distributions were highly positively skewed with a mean and variance indicating overdispersed data. Therefore, negative binomial models were used to compare OSP count and % OSP using SAS GENMODE (v. 9.4). Data are presented as Means ± Standard Error.RESULTSMain effects for Age and Condition were detected in the ELISAs. Plasma IL‐1α did not differ; however, IL‐1β was higher (p&lt;0.05) in Old (30.8±4.6 pg/mL) vs. Young (15.0±4.3 pg/mL). For Condition, IL‐1RA+BMA (49.6±14.1 pg/mL) had higher (p&lt;0.05) plasma IL‐1α vs. CON (7.5E‐15±12.7 pg/mL) and IL‐1RA (9.4±13.2 pg/mL), and both CON (31.7±6.1 pg/mL) and IL‐1RA (41.2±6.3 pg/mL) had higher plasma IL‐1β vs. BMA (6.5±5.8 pg/mL) and IL‐1RA+BMA (12.3±6.8 pg/mL). For the OSP, Age x Condition interactions were detected. OSP count was higher (p&lt;0.05) in Old IL‐1RA+BMA (3977±1631) vs. all other groups except Old IL‐1RA (1422±476). In addition, OSP count was higher (p&lt;0.05) in Old IL‐1RA vs. Young IL‐1RA (212±67), Young BMA (276±80), and Young IL‐1RA+BMA (298±106). OSP count did not differ between Young CON (399±134) and Old CON (561±199). Analysis of % OSP revealed a main effect for diameter. Of the total in circulation, 38.4±7.8% were 15‐30µm, higher (p&lt;0.05) than those 31‐45µm (5.7±1.2%) and 46‐60µm (2.2±0.5%). In addition, the % OSP 0‐14 µm (25.7±3.8%) were higher (p&lt;0.05) vs. 31‐45µm and 45‐60µm. Further, the % OSP 31‐45µm was higher (p&lt;0.05) vs. 46‐60µm.CONCLUSIONPlasma concentrations of IL‐1α and IL‐1β varied with age and condition. Treatment with the IL‐1RA in old rats exacerbated the number of OSP in circulation, with the combined treatment (i.e., IL‐1RA+BMA) having the most dramatic effects. The majority of circulating OSP were 15‐30 µm in diameter, i.e., diameters amenable to embolism.

Impact of adolescent intermittent ethanol exposure on interneurons and their surrounding perineuronal nets in adulthood.

Binge alcohol exposure during adolescence results in long-lasting alterations in the brain and behavior. For example, adolescent intermittent ethanol (AIE) exposure in rodents results in long-term loss of functional connectivity among prefrontal cortex (PFC) and striatal regions as well as a variety of neurochemical, molecular, and epigenetic alterations. Interneurons in the PFC and striatum play critical roles in behavioral flexibility and functional connectivity. For example, parvalbumin (PV) interneurons are known to contribute to neural synchrony and cholinergic interneurons contribute to strategy selection. Furthermore, extracellular perineuronal nets (PNNs) that surround some interneurons, particularly PV+ interneurons, further regulate cellular plasticity. The effect of AIE exposure on the expression of these markers within the PFC is not well understood. The present study tested the hypothesis that AIE exposure reduces the expression of PV+ and choline acetyltransferase (ChAT)+ interneurons in the adult PFC and striatum and increases the related expression of PNNs (marked by binding of Wisteria floribunda agglutinin lectin) in adulthood. Male rats were exposed to AIE (5g/kg/day, 2-days-on/2-days-off, i.e., P25 to P54) or water (CON), and brain tissue was harvested in adulthood (>P80). Immunohistochemistry and co-immunofluorescence were used to assess the expression of ChAT, PV, and PNNs within the adult PFC and striatum following AIE exposure. ChAT and PV interneuron densities in the striatum and PFC were unchanged after AIE exposure. However, PNN density in the PFC of AIE-exposed rats was greater than in CON rats. Moreover, significantly more PV neurons were surrounded by PNNs in AIE-exposed subjects than controls in both PFC subregions assessed: orbitofrontal cortex (CON=34%; AIE=40%) and medial PFC (CON=10%; AIE=14%). These findings indicate that, following AIE exposure, PV interneuron expression in the adult PFC and striatum is unaltered, while PNNs surrounding these neurons are increased. This increase in PNNs may restrict the plasticity of the ensheathed neurons, thereby contributing to impaired microcircuitry in frontostriatal connectivity and related behavioral impairments.

Nephro Defensive Efficiency of Cichorium Intybus Against Toxicity Caused by Copper Oxide Nanoparticles

The penetration of nanotechnology into the medical world is constantly growing, and this raises concerns about its toxicity. This study aimed to illustrate the defense ability of Cichorium intybus (CI) against the nephrotoxicity induced by copper oxide nanoparticles (CuONPs) in a rat model. 32 laboratory rats were evenly distributed into 4 groups, each containing 8 animals. CON rats did not receive any dosage, whereas, CuONPs animals were dosed with copper oxide nanoparticles. As for the CuONPs + CI group, intoxicated rats were dosed with Cichorium intybus extract. Lastly, CI rats were given C.intybus extract only. The parameters of renal function, oxidative stress, and renal histological changes were measured. Animals treated with CuONPs showed negative changes in all examined parameters of kidneys compared to the CON group. However, co-administration of plant extract with NPs showed a clear improvement effect of these toxic changes. We concluded that the high dose of CuONPs causes significant kidney damage, and CI extract has a defensive effect against this toxicity. Keywords: Nephrotoxicity, nanoparticles, oxidative stress, histological changes.

Adolescent Intermittent Ethanol (AIE) Enhances the Dopaminergic Response to Ethanol within the Mesolimbic Pathway during Adulthood: Alterations in Cholinergic/Dopaminergic Genes Expression in the Nucleus Accumbens Shell.

A consistent preclinical finding is that exposure to alcohol during adolescence produces a persistent hyperdopaminergic state during adulthood. The current experiments determine that effects of Adolescent Intermittent Ethanol (AIE) on the adult neurochemical response to EtOH administered directly into the mesolimbic dopamine system, alterations in dendritic spine and gene expression within the nucleus accumbens shell (AcbSh), and if treatment with the HDACII inhibitor TSA could normalize the consequences of AIE. Rats were exposed to the AIE (4 g/kg ig; 3 days a week) or water (CON) during adolescence, and all testing occurred during adulthood. CON and AIE rats were microinjected with EtOH directly into the posterior VTA and dopamine and glutamate levels were recorded in the AcbSh. Separate groups of AIE and CON rats were sacrificed during adulthood and Taqman arrays and dendritic spine morphology assessments were performed. The data indicated that exposure to AIE resulted in a significant leftward and upward shift in the dose-response curve for an increase in dopamine in the AcbSh following EtOH microinjection into the posterior VTA. Taqman array indicated that AIE exposure affected the expression of target genes (Chrna7, Impact, Chrna5). The data indicated no alterations in dendritic spine morphology in the AcbSh or any alteration in AIE effects by TSA administration. Binge-like EtOH exposure during adolescence enhances the response to acute ethanol challenge in adulthood, demonstrating that AIE produces a hyperdopaminergic mesolimbic system in both male and female Wistar rats. The neuroadaptations induced by AIE in the AcbSh could be part of the biological basis of the observed negative consequences of adolescent binge-like alcohol exposure on adult drug self-administration behaviors.

Telmisartan induces a specific gut microbiota signature which may mediate its antiobesity effect

Telmisartan prevents diet-induced obesity (DIO) in rodents. Given that the precise underlying mechanism is not known, we examined whether a gut-related mechanism might be involved. Sprague-Dawley rats received cafeteria diet (CD) for 3 months to develop DIO and were administered either telmisartan (8 mg/kgbw) or vehicle. In addition, pair-fed (PF) rats received CD adjusted to TEL and control rats (CON) only received chow. Stool samples were analysed by 16 S rRNA gene amplicon sequencing. CD-fed rats became obese while TEL, PF and CON rats remained lean. Alpha diversity analyses indicated that bacterial diversity was similar before the study but changed over time. Beta diversity revealed a time-, CD- and telmisartan-dependent effect. The Firmicutes/Bacteroidetes ratio and the abundance of Blautia, Allobaculum and Parasutterella were higher in DIO and PF than in CON, but not in TEL. Enterotype (ET)-like clustering analyses, Kleinberg’s hub network scoring and random forest analyses also indicated that telmisartan induced a specific signature of gut microbiota. In response to stool transfer from telmisartan-pre-treated donor to high-fat fed acceptor mice, body weight gain was slightly attenuated. We attribute the anti-obesity action of telmisartan treatment to diet-independent alterations in gut microbiota as the microbiota from telmisartan-treated, CD-fed rats clearly differed from those of DIO and PF rats. ET-like clustering network, random forest classification and the higher stability in bacterial co-occurrence network analyses indicate that there is more than one indicator species for telmisartan’s specific signature, which is further strengthened by the fact that we cannot identify a single indicator species.

Cerebrospinal fluid proteomics reveal potential protein targets of JiaWeiSiNiSan in preventing chronic psychological stress damage

Context Chinese herbal formula JiaWeiSiNiSan (JWSNS) has been widely used to prevent stress-induced neuropsychiatric ailments in clinics and proven to have therapeutic anti-stress effects on rats. However, the mechanism remains unclear. Objective Based on the proteomics of cerebrospinal fluid (CSF), this study explores the possible mechanism and target proteins of JiaWeiSiNiSan raising stress resilience and preventing stress damage. Materials and methods A 6-week Chronic Unpredictable Mild Stress (CUMS) model was applied on adult Wistar male rats to observe the effects of JWSNS on improving mental stress resilience. Tandem Mass Tag (TMT) proteomics and bioinformatics analysis were used to screen and analyze differentially expressed proteins (DEPs) in CSF. Parallel Reaction Monitoring (PRM) was used to validate target DEPs. Results Significantly decreased sucrose preference, locomotion activity level and accuracy of T-maze, as well as increased immobility time, were observed in CUMS rats compared to CON rats while JWSNS improved above depression-like behaviours. The quantitative proteomics and bioinformatics analysis showed that JWSNS decreased the expression of Rps4x, HSP90AA1, Rps12, Uba1, Rsp14, Tuba1b in CUMS rats CSF (p < 0.05, FDR < 0.5). Immunofluorescence results showed that the number of BrdU/DCX positive cells (p < 0.01) and the relative number of neurons (p < 0.01) in the hippocampus dentate gyrus (DG) of the JSWNS group significantly increased, compared with the CUMS group. Conclusions JWSNS could increase mental stress resilience and prevent stress damage by regulating proteins in CSF. This study provides a scientific basis for further study on Chinese formulas preventing mental illness.

Autoimmune activation of the GnRH receptor induces insulin resistance independent of obesity in a female rat model.

Polycystic ovary syndrome (PCOS), a metabolic and reproductive disease, is frequently associated with type 2 diabetes. We have demonstrated activating autoantibodies (AAb) directed toward the second extracellular loop (ECL2) of the gonadotropin‐releasing hormone receptor (GnRHR) are present in a significant subgroup of PCOS patients. It is unclear whether GnRHR‐AAb can induce peripheral tissue insulin resistance (IR) in animal models. Sixteen rats were divided equally into a GnRHR ECL2 peptide‐immunized group (IMM group) and a control group (CON group). Sera GnRHR‐AAb titer, luteinizing hormone (LH), and testosterone (T) were higher in IMM rats compared with CON rats. No significant difference in fasting blood glucose was observed between the two groups. However, the plasma glucose level at other time points of the IMM group was higher than that of the CON group during an intraperitoneal glucose tolerance test (IPGTT) and an insulin tolerance test (ITT) (p < 0.01). These data support the likelihood of the GnRHR‐AAb induction of glucose intolerance and IR. Compared with the CON group, the IMM group showed a significant increase in insulin‐stimulated phosphorylation of IRS‐1 (p‐IRS‐1 S636/639) and a decrease in insulin‐stimulated phosphorylation of Akt (p‐AKT S473). Expression of the glucose transport genes including GLUT‐2 in liver and GLUT‐4 in white adipose tissue and skeletal muscle was significantly decreased in IMM rats compared with the CON rats. Serum levels of proinflammatory cytokines (TNF‐α, IL‐1α, and IL‐18) were increased, while anti‐inflammatory cytokines (IL‐4 and IL‐10) were decreased in the IMM group. Taken together, elevated GnRHR‐AAb enhanced LH, hyperandrogenism, and inflammation. These changes are likely related to the observed peripheral tissue IR through the downregulation of the insulin‐stimulated IRS/PI3K/Akt/Glut signaling pathway.

Simulated vehicle exhaust exposure (SVEE) in rats impairs renal mitochondrial function

ABSTRACT Purpose Vehicle exhaust emissions primarily comprise of nitrogen, oxygen, water, CO2, NO2, CO, hydrocarbons and particulate matter. While adverse effects of hydrocarbon and particulate matter on cardiovascular functions are known, the effect of pro-oxidants CO2, NO2 and CO are not clear. Methods Here, using an animal model of a simulated mixture of pro-oxidants (0.04% CO2, 0.9 ppm NO2 and 3 ppm CO with air as a base), we examined the effect of simulated vehicle exhaust exposure (SVEE) on various cardiovascular parameters. Male Sprague-Dawley rats were exposed to SVEE or ambient air (Control: CON) for 30 min/day for 2 weeks. Thereafter, systolic and diastolic blood pressure, heart rate and glomerular filtration rate were measured. Later, rats were sacrificed, blood plasma and kidneys were collected. Results The systolic and diastolic blood pressure, heart rate and glomerular filtration rate remained unchanged. Plasma corticosterone increased in SVEE rats when compared to CON group. Plasma 8-isoprostane, a systemic marker of oxidative stress, increased while total antioxidant capacity decreased in SVEE but not in CON. Kidney cortical tissue homogenates exhibited increase in superoxide, hydrogen peroxide and protein carbonylation in SVEE but not CON, all indicative of heightened oxidative stress. Renal cortical mitochondrial SOD activity was significantly reduced in SVEE than CON. Conclusion Significant decline in mitochondrial respiration and oxygen consumption was observed, in addition to low ATP, reduced ATP synthase and cytochrome C oxidase levels, as well as accelerated mitochondrial fission, and reduced fusion processes, were observed in SVEE than CON rats, all indicative of renal mitochondrial impairment.

Influence of Obesity on Vascular Dysfunction Following Hemorrhagic Shock

Obesity is associated with higher rates for complications after traumatic hemorrhage, including higher incidence of multiple organ failure, thrombosis, and mortality. These adverse events are associated with prolonged tissue hypoperfusion, hypercoagulability, and edema that reflect profound vascular damage. However, the combined impact of obesity and traumatic hemorrhage on vascular function is not well understood. Therefore, we sought to determine the impact of diet‐induced obesity on vasoconstrictor and vasodilator properties of arteries before and after hemorrhagic shock.Male Sprague‐Dawley rats were fed a control (CON; 13.5% kcal fat, n=10), moderate fat (MF, 33% kcal fat n=9) or high fat (HF, 60% kcal fat, n=10) diet for 6–8 weeks. Hemorrhagic shock was induced by laceration of a mesentery artery and a Grade V splenic injury under anesthesia, and a period of hypotension (mean arterial pressure = 30–40 mm Hg) was maintained for 30 minutes. We assessed vasoconstrictor and vasodilator properties in isolated mesenteric arteries harvested before and after hemorrhagic shock. Before hemorrhage, peak norepinephrine (NE) vasoconstriction was higher in CON rats (78±4%) versus HF (62±5%, p=0.01), while MF rats exhibited a peak NE vasoconstriction not different from either groups (71±3%, p&gt;0.05 vs CON and HF). Baseline endothelium‐dependent dilation, assessed as the maximal response to acetylcholine, was not different between groups (CON: 97±1%, MF: 88±4%, HF: 92±3%, p=0.08). Hemorrhage lowered the peak NE vasoconstriction (post: 61±8%, p=0.05) and endothelium‐dependent dilation in CON rats (post: 90±2%, p&lt;0.001), but had no effect on NE vasoconstriction and endothelium‐dependent dilation in MF and HF rats (p&gt;0.05 vs pre‐hemorrhage). Endothelium‐independent dilation, assessed as the response to sodium nitroprusside, was similar pre‐ and post‐hemorrhage in all groups (p&gt;0.05).In conclusion, we find traumatic hemorrhage induces vascular dysfunction characterized by a reduced vasoconstrictor response and impaired endothelial function, but only in rats fed a normal chow diet. While obesity results in vascular dysfunction at baseline, it does not contribute to a further worsening of function after traumatic hemorrhage.Support or Funding InformationFunded by a UO‐OHSU Collaborative Seed Grant

Modulation effects of black-vinegar-based supplement against a high-fat dietary habit: antiobesity/hypolipidemic, antioxidative, and energy-metabolism effects.

An imbalanced fat or excess energy intake always results in obesity and increased serum/liver lipids, thus leading to metabolic syndromes. Given the bioactive components in black vinegar (BV), such as branched amino acids, phenolic profile, and mineral contents, we investigated the antiobesity effects of BV-based supplements in rats fed a high-fat diet (HFD). HFD (30% fat, w/w) feeding increased (P < 0.05) body weight, weight gains, weights of livers and mesenteric, epididymal, and perirenal adipose tissues, and serum/liver triglyceride levels relative to those of rats fed a normal diet (4% fat, w/w; CON). These increased values were ameliorated (P < 0.05) by supplementing with BV-based supplements but were still higher (P < 0.05) than those of CON rats. The increased areas of perirenal adipocytes in rats fed with an HFD were also decreased (P < 0.05) by supplementing with BV-based supplements, which might result from an upregulation (P < 0.05) of 5'-adenosine monophosphate-activated protein kinase (AMPK), carnitine palmitoyltransferase-1 (CPT1), and uncoupling protein-2 (UCP2) in the perirenal adipose tissues. A similar effect was observed for AMPK, peroxisome proliferator-activated receptor alpha, retinoid X receptor alpha, CPT1, and UCP2 gene and protein levels in livers (P < 0.05). Generally, BV-based supplements increased the fecal triglyceride, cholesterol, and bile acid levels of rats fed with an HFD, which partially contribute to the lipid-lowering effects. Furthermore, BV-based supplements increased (P < 0.05) hepatic Trolox equivalent antioxidant capacity and lowered (P < 0.05) serum/liver thiobarbituric acid reactive substances values in HFD-fed rats. In a chronic high-fat dietary habit, the food-grade BV-based supplement is a good daily choice to ameliorate obesity and its associated comorbidities. © 2020 Society of Chemical Industry.