Single Intraperitoneal Injection Research Articles

ATP-Gated P2X7-Ion Channel on Kidney-Resident Natural Killer T Cells and Memory T Cells in Intrarenal Inflammation.

The P2X7 ion channel, a key sensor of sterile inflammation, has been implicated as a therapeutic target in glomerulonephritis, and P2X7-antagonistic nanobodies can attenuate experimental glomerulonephritis. However, little is known about the expression of P2X7 on renal immune cells. We used conventional immunofluorescence of kidney sections as well as intraperitoneal injection of nanobodies in mice followed by flow cytometry analysis of parenchymal T cells and RNA-sequencing to elucidate the expression and function of P2X7 on parenchymal and vascular immune cells in the mouse kidney. Our study showed that parenchymal T cells, including a large subset of natural killer T cells and CD69+ tissue-resident memory T cells, display much higher cell surface levels of P2X7 than vascular T cells. After a single intraperitoneal injection of P2X7-blocking nanobodies, P2X7 on parenchymal T cells was fully occupied by the injected nanobodies within 30 min. This resulted in an effective protection of these cells from NAD-induced cell death during cell preparation. Conversely, systemic injection of NAD that mimics sterile inflammation results in the selective depletion of P2X7hiCD69hi T cells from the kidney parenchyma. Our study uncovered a novel purinergic regulatory mechanism affecting kidney-resident T cell populations.

Co-Administration of Hibiscus sabdariffa linn and Daflon-500® mitigates weight loss, hyperglycemia, hematological and oxidative changes in male rats with type-2 diabetes

BackgroundDiabetes mellitus is a complex metabolic disorder associated with persistent hyperglycemia and metabolic imbalance. Flavonoid compounds possess notable qualities in mitigating diabetes and its complications. AimHibiscus sabdariffa linn and daflon-500® were assessed for their effects on body weight, blood glucose/indices, and oxidative changes in type-2 diabetes rats over 28 days. Methods35 male rats were randomly divided into seven groups (n=5). Diabetes mellitus was induced in groups 2-7 via a single intraperitoneal injection of streptozotocin (STZ; 55mg/kg). Group 1(DW) received distilled water (2mL/kg) only. Group 2 (STZ; untreated), group 3 {MET; metformin (200mg/kg)}, group 4 {HS; Hibiscus sabdariffa (400mg/kg)}, group 5 {DAF; daflon-500® (1000mg/kg)}, group 6 {DAF+HS; daflon-500® (1000mg/kg) and Hibiscus sabdariffa (400mg/kg)}, group 7 {DAF+MET+HS; daflon-500® (1000mg/kg), metformin (200mg/kg), and Hibiscus sabdariffa (400mg/kg)}. Body weight and blood glucose were assessed fortnightly. Blood and hepatic/pancreatic tissues were collected for hematological and antioxidant testing. ResultsGroups 2-7 revealed a significant (P<0.05) decrease in body weight on day 14 compared to the increased growth rate observed on day 28 (Group 7).Blood glucose levels significantly increased on days 1 and 14 in groups 2-7 but declined on day 28 in groups 4-7. Significantly higher packed cell volume, hemoglobin levels, red blood cell counts, lymphocytes, and mean corpuscular hemoglobin were observed in groups 5-7. Platelet counts were higher in Groups 2 and 5. Untreated group 2 exhibited considerably reduced catalase activity, glutathione, and malondialdehyde concentrations compared to the treated groups. ConclusionHibiscus sabdariffa linn and Daflon-500® expressed substantial hypoglycemic, antioxidant, and hemato-protective effects.

Genetic machinery which accompanies metaplasticity operates differentially in experimental model of autism

AbstractThe present study investigated metaplasticity‐related mRNA expressions in valproic acid (VPA)‐rats, focusing on the PI3K/AKT pathway. Wistar dams were treated with a single intraperitoneal injection of 600 mg/kg VPA or saline on embryonic day E12.5 or an equal volume of saline solution. Three behavioral tests were conducted on these males' offspring: grid‐walking test, negative geotaxis test, and three‐chamber social interaction test. Metaplasticity was induced in 60‐day‐old male progeny by giving high‐frequency stimulation for 5 minutes following low‐frequency stimulation to the perforant pathway. For the baseline stimulation protocol (n = 6), stimulation was delivered to the dentate gyrus at the previously determined stimulation intensity (0.33 Hz 0.175 msec 30 s) for 75 min. The percent change of slope of field excitatory postsynaptic potential (fEPSP) and amplitude of population spike were calculated 55–60 min after induction protocol. The mRNA levels of PI3K, PTEN, AKT, GSK‐3β, and MAPT were measured in the hippocampus by using quantitative rt‐PCR. We found that offspring of VPA‐treated rats showed significantly impaired sensorimotor coordination, decreased sociability, impaired preference for social novelty, and reduced input–output curve of fEPSP slope, compared to control animals. Despite a similar metaplastic response, mRNA levels of genes of interest were similar but considerably down‐regulated after induction in offspring of VPA‐treated dams. Our study provides evidence that the induced expression of autism‐related genes has evolved to enable an adaptation mechanism during metaplastic control of long‐term potentiation.

Integrating network pharmacology and in vivo pharmacological investigation for deciphering the mechanism of Simiao Yong’an decoction in alleviating rheumatoid arthritis

Objective: To investigate the mechanism of Simiao Yong’an Decoction in alleviating the damage of rheumatoid arthritis (RA), and accordingly provide pharmacological evidence and further experimental support for the application of Simiao Yong’an Decoction, as well as provide a reference direction for the development of new drugs for RA treatments. Materials and methods: Arthritis was induced in Balb/c mice. The animals were randomly divided into 4 groups: control, model, methotrexate (MTX), and Simiao Yong’an Decoction group (n = 6). The Simiao Yong’an Decoction group was orally administered 4.5 g·kg−1 of Simiao Yong’an Decoction every day, and the MTX group received a single intraperitoneal injection of 2 mg·kg−1 of MTX. The arthritis symptoms of joints were scored every day after injecting a cocktail antibody. The histopathology injuries and bone destruction of joints were measured by hematoxylin and eosin staining, Safranin-O fast green staining, tartrate-resistant acid phosphatase staining, and micro–computed tomography. The regulation of Simiao Yong’an Decoction on leukocytes was analyzed by flow cytometry. The expression of inflammatory cytokines, chemokines, and matrix metalloproteinases in joints was detected by quantitative real-time polymerase chain reaction. In the network pharmacology analysis, drug targets of Simiao Yong’an Decoction were obtained from Swiss Target Prediction. Disease targets of RA were obtained from the Therapeutic Target Database and Disgenet. The ingredients-targets network, protein-protein interaction network, and gene ontology function enrichment were performed and visualized by Cytoscape software. Results: In the collagen antibody-induced arthritis (CAIA) murine model, Simiao Yong’an Decoction effectively improved arthritis symptoms, decreased the expression of inflammatory cytokines, and reduced joint pathologic damage and bone destruction. In the network pharmacology analysis, multiple active ingredients of Simiao Yong’an Decoction were found to regulate genes related to inflammation, chemotaxis, and collagen degradation in RA, and reduce RA damage by regulating biological processes including leukocyte migration, myeloid cell homeostasis, and collagen degradation metabolism. We further verified that Simiao Yong’an Decoction effectively regulated the number of myeloid cells in CAIA mice. Moreover, Simiao Yong’an Decoction suppressed the expression of myeloid cell-associated chemokines and matrix metalloproteinases in inflamed joints of CAIA mice. Conclusion: Simiao Yong’an Decoction effectively reduces RA damage by regulating myeloid cells and reducing collagen degradation.

Hypoglycemic and antihyperglycemic effects of Abelmoschus esculentus and Alchornea cordifolia in normal and alloxan-induced diabetic rats

BackgroundAbelmoschus esculentus and Alchornea cordifolia are commonly used in Traditional Chinese Medicine (TCM) to treat several diseases. Abelmoschus esculentus is used to treat infertility and menorrhagia, while Alchornea cordifolia is used for the treatment of venereal diseases, cough, and diarrhoea. However, very few studies assessed the antidiabetic effects of these plants. Therefore, this study aimed to investigate the hypoglycemic and antihyperglycemic effects of aqueous extract of A. esculentus fruits and A. cordifolia leaves. Material and MethodsFresh Abelmoschus esculentus fruits and the powder from the dried leaves of Alchornea cordifolia leaves were prepared by maceration in the aqueous phase (200g/100mL and 50g/100mL respectively) for 24 hours, then filtered and concentrated in an oven at 45°C. Diabete was induced to male Wistar rats by a single intraperitoneal injection of alloxan (150 mg/kg, b.w). Rats with a blood glucose level greater than or equal to 200 mg/dL were selected, divided into groups and were daily administered orally with either aqueous extracts of A. esculentus at 30 mg/kg (AEAE30) or A. cordifolia at 400 mg/kg (AEAC400) for 14 consecutive days. For comparison, acarbose (100 mg/kg), glibenclamide (5 mg/kg), and metformin (500 mg/kg) were administered orally as reference drugs. Moreover, insuline was also used as a positive control and administered intraperitoneally at a dose of 5 IU/kg. Then, blood glucose levels, oral glucose tolerance test, oral maltose tolerance club, body weight and hemoglobin were assessed. For evaluation of the aqueous extracts in the intestinal transit, imodium (2 mg/kg, p.o) and fructine (5 mg/kg, p.o) were used as a positive control to determine the spasmolytic and laxative activities, respectively. The histopathological study of the liver, kidney, pancreas, testis, epididymis, and seminal vesicle was also carried out using the hematoxyline & eosin (H&E) technique. ResultsAEAE30 and AEAC400 significantly reduced (P<0.001) fasting blood glucose (FBG) levels and significantly prevented (P<0.001) postprandial glycemia in AI-db rats following oral glucose tolerance test (OGTT) and oral maltose tolerance test (OMTT) in alloxan-induced diabetic rats (AI-db). In normoglycemic and insulin-resistant (IR) rats, AEAE30 significantly prevented the post-prandial blood glucose level during the OGTT (P<0.01) only in normoglycemic rats. At the end of treatment, AEAE30 significantly reduced the relative weight of the liver (P<0.01) and significantly increased (P<0.001) the relative weight of the testes and pancreas while AEAC400 significantly increased the relative weight of the testes compared to untreated AI-db rats. The histopathological study revealed a restoration of alloxan-induced tissue damage close to the normal control group in AI-db animals treated with plant extracts, as well as an increase in sperm density in the epididymis unlike in the untreated AI-db group. ConclusionThese findings suggested that AEAE and AEAC exhibited hypoglycemic and antihyperglycemic activities and could be therefore a useful source of antidiabetic agent.

Antihypoxic effect of a combination of bemethyl and thymoquinone under hypercapnic hypoxia

The antihypoxic effect of a combination of bemethyl and thymoquinone after single intraperitoneal injection was studied in a model of hypercapnic hypoxia in a hermetic volume in mice. Thymoquinone had a dose-dependent protective effect by increasing the resistance of animals to hypoxia. At a dose of 10 mg/kg, it increased the life expectancy by 75.9 % compared with the control group, and at a dose of 30 mg/kg – by 90.2 %. The average effective dose of thymoquinone calculated by the regression analysis in this model was 14.1 mg/kg and the average effective dose of bemethyl was 148 mg/kg. As part of a combination of bemethyl and thymoquinone in the ratio of 5 : 1, the activity of the compounds increases significantly, ED50 of bemethyl decreases to 39.0 mg/kg, the same indicator for thymoquinone becomes 7.8 mg/kg. Administration of a combination of 100 mg/kg of bemethyl and 20 mg/kg of thymoquinone to mice leads to increasing their lifespan under the standard hypoxic conditions up to 41.2 ± 5.7 minutes, which is 120.5 % more than in animals receiving placebo. The analysis of pharmacodynamics of a combination of drugs according to T. Chou showed an additive type of interaction typical for compounds with different mechanisms of primary action. It was established that a combination of bemethyl and thymoquinone in the weight ratio of 5 : 1 demonstrate synergism, which makes it possible to reduce 1.8–4.0 times the doses of components in their combined use.

Sex-specific NLRP3 activation in neutrophils promotes neutrophil recruitment and NETosis in the murine model of diffuse alveolar hemorrhage.

Diffuse alveolar hemorrhage (DAH) is a life-threatening complication of systemic lupus erythematosus and small vessel vasculitis. We previously showed that neutrophil extracellular traps (NETs) were associated with the pathogenesis of pristane-induced DAH and demonstrated that neutrophil NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome assembly participated in NET generation under sterile stimulation. We investigated whether NLRP3 inflammasome assembly in neutrophils may drive pulmonary NETosis in a mouse model of pristane-induced DAH. C57BL/6J mice received a single intraperitoneal injection of 0.5mL of pristane. Neutrophil NLRP3 inflammasome assembly and NETs were characterized by immunofluorescence staining of apoptosis-associated speck-like protein a CARD (ASC), co-staining of DNA, and citrullinated histones, respectively. Clinical status of mice was assessed 11 days after pristane injection by measurement of arterial oxygen saturation and of weight loss; severity of lung injury was determined using a quantification score from hematoxylin-eosin-stained slides. Pristane induced ASC speck formation in neutrophils and we confirmed that NLRP3 inflammasome was involved in NET generation after pristane stimulation in vitro. NLRP3 deficiency reduced the severity of pristane-induced DAH in female, but not male mice. Interestingly, NLRP3 deficiency reduced the number of neutrophils and NETs in the lungs of females compared to males. Our results suggest a link between female sex-specific NLRP3 inflammasome activation and subsequent pulmonary NETosis in the development of pristane-induced DAH. Therefore, we identified NLRP3 inflammasome as a potential new therapeutic target in this severe complication of pro-female autoimmune disease for which specific inhibitors of NLRP3 are currently developed.

Cinnamon Polyphenols Ameliorate the Dysregulation of Cardiac Energy and Autophagic Homeostasis in Rats with Diabetic Cardiomyopathy via the mTOR/PGC1α Pathway

Background Inflammation mediated by hyperglycemia, lipid metabolism disorders, abnormal myocardial energy metabolism, myocardial cell damage, and changes in ventricular structure are important mechanisms in the occurrence and development of diabetic cardiomyopathy (DCM). Cinnamon polyphenols (CP) are extracted from the traditional Chinese medicine cinnamon, which is believed to have cardioprotective effects and has been used in China for hundreds of years. Purpose This study aimed to investigate the therapeutic effects of CP on DCM and its possible mechanisms. Methods The DCM model was established by a single intraperitoneal injection of 85 mg/kg 1% streptozotocin in rats. The treatment group was orally administered 135 mg/kg CP for 28 consecutive days, a dosage verified by experiments on AC16 myocardial cells and detecting liver and kidney injury markers. Results CP could reduce high blood sugar, high blood lipids, left ventricular mass index, levels of myocardial injury markers, adenosine diphosphate (ADP), and cyclic adenosine monophosphate (cAMP) in the model group. It also improved weight loss, and increased myocardial adenosine triphosphate (ATP), adenosine monophosphate (AMP), phosphocreatine (PCr), and ATP/ADP, AMP/ATP, and PCr/ATP ratios. The expression of p-mTOR/mammalian target of rapamycin (mTOR), p62, and peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α) in the myocardium of the CP group was enhanced, while the expression of LC3II/LC3I was inhibited. Conclusion CPs can alleviate myocardial damage by reducing blood sugar, blood lipids, and abnormal autophagy levels while promoting the restoration of myocardial energy metabolism homeostasis.

The Glucagon-Like Peptide-1 (GLP-1) Receptor Agonist Liraglutide Regulates Sirtuin-1-Mediated Neutrophil Extracellular Traps to Improve Diabetes-Induced Bone Metabolism Imbalance

Background: Diabetes mellitus (DM) is a chronic metabolic disorder that disrupts normal bone remodeling. Objectives: This study aimed to investigate how the glucagon-like peptide-1 (GLP-1) receptor agonist liraglutide (LIR) addresses bone metabolism imbalances induced by type-II diabetes. Methods: Type-II diabetic rat models were established through a single intraperitoneal injection of streptozotocin (STZ). Blood glucose levels were measured using a blood glucose meter, and insulin levels were assessed using an assay kit. Bone formation markers [alkaline phosphatase (ALP), osteocalcin (OCN), and procollagen I N-terminal propeptide (PINP)] and bone resorption markers [tartrate-resistant acid phosphatase (TRACP) and CTX-1] were monitored using assay kits. Bone marrow mesenchymal stem cells (BMSCs) were cultured in vitro under high-fat and high-glucose (HFHS) conditions to mimic diabetic bone metabolism dysregulation. Neutrophil extracellular traps (NETs) formation was examined through immunofluorescent staining and Western blot analysis. Results: Liraglutide was found to reduce STZ-induced NETs formation, as indicated by decreased expression of cit-H3 by 36.90% - 53.57%, myeloperoxidase (MPO) by 55.81% - 65.12%, NE by 53.95% - 65.17%, and PAD4 by 46.81% - 63.83%, alongside increased Sirtuin-1 (SIRT1) expression in femur tissue (70.71% - 91.19%). In vitro, LIR enhanced osteogenesis and inhibited apoptosis, effects that were partially reversed by SIRT1 knockdown. Additionally, SIRT1 knockdown partially restored LIR-induced reductions in oxidative stress, inflammation, and NETs formation. Conclusions: LIR mitigates diabetes-induced bone metabolism imbalance by inhibiting NETs formation through SIRT1 mediation.

Biochemical Studies of Averrhoa carambola Ethanol Leaf and Fruit Extracts on Alloxan-induced Diabetic Rats

The use of herbal medicines has always been an option to treat diseases such as diabetes cancer and other ailments. This study aimed to evaluate the effect of Averrhoa carambola leaf and its fruit ethanol extracts on biochemical parameters and histopathology features. Forty five albino rats were assigned into 9 groups of 5 rats each. Except for the normal control group, others were induced to be diabetic by a single intraperitoneal injection of alloxan. Group 1 (normal control) and Group 2 (diabetic untreated) received water. Group 3 received 2.5mg/kg glibenclamide, Groups 4-9 received graded doses (250mg/kg, 500mg/kg and 750mg/kg) of leaf and fruit extracts of Averrhoa carambola. Treatment lasted 14days. At the end of the treatment, blood samples and organ (pancreas) were collected from the animals for biochemical and histopathological evaluation. Treatment of alloxan-induced diabetic rats significantly (p&lt;0.05) reduced the glucose concentration across all treated groups. Following induction of diabetic mellitus, the serum activities of liver enzymes (Alanine amino transferase, Aspartate aminotransferase and Alkaline phosphatase), low-density lipoprotein cholesterol (LDL-C) and triglyceride were significantly (p&lt;0.05) elevated in diabetic untreated group with corresponding reduction in serum protein, albumin and high-density lipoprotein cholesterol (HDL-C) compared to the normal control and treated groups. However, the fruit extracts exhibited a high increase in urea and creatinine levels especially at dose of 750mg/kg body weight when compared with the leaf though not significantly higher than the diabetic untreated group. From histopathology studies, the groups treated with the leaf extract of Averrhoa carambola showed greater structural improvement and an increase in pancreatic islet cells. Therefore this study showed that although ethanol extracts of Averrhoa carambola leaf and fruit have antidiabetic properties, the leaf extract possesses more antidiabetic effect.

HISTOLOGICAL CHANGES IN THE RAT SPLEEN IN THE EARLY STAGES OF EXPERIMENTAL STREPTOZOTOCIN-INDUCED DIABE-TES MELLITUS

Diabetes is a systemic disorder that has a significant impact on a number of different organ systems within the body. Studying changes in the spleen in diabetic rats aids in understanding these changes and their effect on immune function. The objective of the study is to investigate the characteristics of morphological changes in the structural components of the spleen in the early stage of experimental diabetes.Results. Morphological changes in the spleen during the early stages of streptozotocin-induced experimental diabetes mellitus were observed. Diabetes was induced by a single intraperitoneal injection of streptozotocin. Histological examination of the spleen was conducted 14 and 28 days after diabetes induction. In healthy rats, the spleen exhibited a normal structure with a clear distinction between red and white pulp. Morphometric analysis showed that lymphoid nodules had a diameter of (445.92±12.6) μm and a marginal zone thickness of (76.22±5.12) μm. After 14 days of experimental diabetes, histological preparations revealed reduced lymphoid nodules of white pulp with a diameter of (435.83±14.28) μm and germinal centers (139.97±5.74) μm. After 28 days, lymphoid nodules further decreased to (392.37±10.29) μm, and germinal centers - to (125.93±5.71) μm. The width of the periarterial lymphoid sheaths decreased to (67.91±3.62) μm, indicating early changes in T-cell zones.Conclusion. During the initial stages of diabetes mellitus in experimental settings, notable structural modifications occur in the spleen. Specifically, there is a reduction in the size of lymphoid nodules, germinal centers, and various zones of the spleen, as well as a decrease in the density of lymphocytes. These alterations may suggest a severe immunological dysfunction in rats used for experimentation.

Semaglutide Ameliorates Diabetic Neuropathic Pain by Inhibiting Neuroinflammation in the Spinal Cord.

Male Wistar rats, each weighing between 300 and 350 g, were categorized into four groups: one non-diabetic sham group and three diabetic groups. The diabetic group received a single intraperitoneal injection of streptozotocin (STZ) at a dosage of 60 mg/kg to induce diabetic neuropathy. After 4 weeks of STZ injection, one diabetic group was given saline (vehicle), and the other two were treated with either 1× SEMA (1.44 mg/kg, orally) or 2× SEMA (2.88 mg/kg, orally). Following a 4-week course of oral drug treatment, behavioral, biochemical, and immunohistochemical analyses were carried out. The mechanical allodynia, thermal hyperalgesia, blood glucose, advanced glycation end products (AGEs), plasma HbA1C, and spinal inflammatory markers were evaluated. SEMA treatment significantly reduced both allodynia and hyperalgesia in the diabetic group. SEMA therapy had a limited impact on body weight restoration and blood glucose reduction. In diabetic rats, SEMA lowered the amounts of pro-inflammatory cytokines in the spinal cord and dorsal horn. It also lowered the activation of microglia and astrocytes in the dorsal horn. SEMA significantly reduced HbA1c and AGE levels in diabetic rats compared to the sham control group. These results indicate SEMA's neuroprotective benefits against diabetic neuropathic pain, most likely by reducing inflammation and oxidative stress by inhibiting astrocyte and microglial activity. Our findings suggest that we can repurpose GLP-1 agonists as potent anti-hyperalgesic and anti-inflammatory drugs to treat neuropathic pain without serious side effects.

Nephroprotective effects of the soluble guanylyl cyclase stimulator, riociguat in doxorubicin-induced acute kidney injury in rats

This study aimed to investigate the potential protective effects of riociguat, a soluble guanylyl cyclase (sGC) stimulator, on kidney function and structure in rats with acute kidney injury (AKI) induced by the chemotherapeutic drug doxorubicin (DX). Rats were subjected to a single intraperitoneal injection of DX (13.5 mg/kg) on the 5th day, either alone or in combination with low-dose riociguat (3 mg/kg/day), or high-dose riociguat (10 mg/kg/day) for 8 consecutive days. Various markers related to kidney function, oxidative stress, and inflammation were measured in plasma and urine. Kidney tissues were examined histopathologically. DX-induced nephrotoxicity was characterized by increased plasma urea, creatinine, uric acid and neutrophil gelatinase-associated lipocalin (NGAL). DX also decreased creatinine clearance and albumin levels and increased urinary N-acetyl-β-D-glucosaminidase (NAG) activity. Furthermore, DX increased the inflammatory markers interleukin 1 beta (IL-1 β), interleukin 6 (IL-6) and tumor necrosis factor alpha (TNF-α). DX further induced oxidative stress injury evidenced by decreased glutathione reductase (GR) activity, total antioxidant capacity (TAC), superoxide dismutase (SOD) and catalase levels and increased malondialdehyde (MDA) levels. Concomitant treatment with riociguat ameliorated these DX-induced changes with parallel histopathological improvements but the effects were more favorable with high-dose riociguat. The observed renoprotective effects of riociguat can be partly attributed to the anti-inflammatory and anti-oxidant properties of this drug.

Sigma-1 receptor modulation by fluvoxamine ameliorates valproic acid-induced autistic behavior in rats: Involvement of chronic ER stress modulation, enhanced autophagy and M1/M2 microglia polarization

Autism spectrum disorder (ASD) is a neurodevelopmental disorder. While, fluvoxamine (FVX) is an antidepressant and widely prescribed to ASD patients, clinical results are inconclusive and the mechanism of FVX in the management of ASD is unclear. This study determined the potential therapeutic impact of FVX, a sigma-1 receptor (S1R) agonist, against the valproic acid (VPA)-induced model of autism. On gestational day 12.5, Wistar pregnant rats were given a single intraperitoneal (i.p.) injection of either VPA (600 mg/kg) or normal saline (10 mL/kg, vehicle-control). Starting on postnatal day (PND) 21 to PND 50, FVX (30 mg/kg, P·O. daily) and NE-100, (S1R) antagonist, (1 mg/kg, i.p. daily) were given to male pups. Behavior tests and histopathological changes were identified at the end of the experiment. In addition, the cerebellum biomarkers of endoplasmic reticulum (ER) stress and autophagy were assessed. Microglial cell polarization to M1 and M2 phenotypes was also assessed. FVX effectively mitigated the histopathological alterations in the cerebellum caused by VPA. FVX enhanced sociability and stereotypic behaviors in addition to its noteworthy impact on autophagy enhancement, ER stress deterioration, and controlling microglial cell polarization. The current investigation confirmed that the S1R agonist, FVX, can lessen behavioral and neurochemical alterations in the VPA-induced rat model of autism.

Effects of Thymol and Menthol on Diethylnitrosamine-induced Changes in SOX2 Expression and TGF-β/SMAD3 Signaling in Mice’s Liver and Kidneys

Background: The signaling pathway of the transforming growth factor (TGF)-β/SMAD3 plays a crucial role in developing liver and kidney fibrosis. TGF-β is linked to metastasis through SRY-related, high-mobility box (SOX-2). The objective of this study was to investigate the impact of thymol and menthol, two monoterpene compounds, on the SOX-2 expression and TGF-β/SMAD3 signaling in the liver and kidneys of mice after administering diethylnitrosamine (DEN). Methods: The Balb/C male mice, aged 14 days, were administered a single intraperitoneal injection of DEN (25 mg/kg) and then given daily either thymol (10 mg/kg) or menthol (50 mg/kg) for 26 consecutive weeks. The mRNA levels of TGF-β, SMAD3, and SOX-2 were measured after 4 and 26 weeks of treatment. Results: The administration of DEN increased the expressions of TGF-β, SMAD3, and SOX-2 in the liver of animals after 4 and 26 weeks; however, their expressions in the kidneys increased only at the end of the 26th week compared to that of the control group. Thymol and menthol inhibited the expression of TGF-β, SMAD3, and SOX-2 in the liver and kidneys of the mice injected with DEN. Conclusion: Based on the study findings, we concluded that thymol and menthol prevented carcinogenesis induction via expression of DEN in the liver and kidneys by inhibiting the associated expression of epithelial-mesenchymal transition and SOX-2.

Therapeutic Potential of N-acetylcysteine and Glycine in Reducing Pulmonary Injury in Diabetic Rats.

Diabetes mellitus is associated with systemic complications, including the development of pulmonary injury, characterized mainly by excessive accumulation of extracellular matrix components and inflammatory cell infiltration in lung tissue. This process is driven by oxidative stress and chronic inflammation, both caused and exacerbated by hyperglycemia. N-acetylcysteine (NAC) and glycine, known for their antioxidant and anti-inflammatory effects, offer potential therapeutic benefits in mitigating diabetes-induced lung injury. The study aimed to investigate the effects of supplementation by either NAC or glycine or their combination on reducing lung injury in rats with type 1 diabetes Materials and methods: The study used 30 adult Wistar albino rats (10 weeks old, weighing between 180 g and 380 g). Six of them were used as controls, while 24 adult rats (10 weeks old, 180-380 g) with type 1 diabetes, induced through a single intraperitoneal injection of streptozotocin (STZ) at a dose of 55 mg/kg, were randomly assigned to four experimental groups: control (CTL), diabetic (Db), NAC treatment (diabetic+NAC), glycine treatment (diabetic+glycine), and combined NAC and glycine treatment (diabetic+NAC+glycine). NAC (100 mg/kg) and glycine (250 mg/kg) were administered orally for 12 weeks. At the end of the study, lung tissues were collected for histopathological examination. Qualitative, semi-quantitative, and stereological histological analysis was used to analyze structural changes in the lung tissue. Semi-quantitative scoring was carried out to evaluate the extent of inflammation, while stereological analysis was performed to determine the volume density of alveolar spaces and septal connective tissue. The semi-quantitative scoring included scores ranging from 0 (absent), 1 (minimal), 2 (mild), 3 (moderate), to 4 (severe). Qualitative histological analysis revealed pronounced inflammation and fibrosis in the lungs of untreated diabetic rats, characterized by thickened alveolar septa and immune cell infiltration. Both treatments with NAC and glycine individually reduced inflammation and fibrosis compared to untreated diabetic rats. The greatest improvement was observed in the NAC+glycine group, where the alveolar structure appeared almost normal, with minimal inflammation. Semiquantitative analysis showed statistically significant differences in peribronchial and peribrochiolar infiltrates between the diabetic group (2.16±0.47) and the control group (0.33±0.21, p=0.026). The combination of NAC and glycine significantly reduced peribronchial and peribronchiolar infiltrates (0.33±0.33, p=0.026) compared to the diabetic group. Similarly, septal inflammatory infiltrates were significantly lower in the NAC+glycine group (1±0.36) compared to diabetic rats (3.33±0.33, p=0.004). Total airway inflammatory infiltration was also significantly reduced in the NAC+glycine group (1.33±0.33, p=0.002) compared to the diabetic group (5.5±0.5). As the combination of NAC and glycine demonstrated protective effects against lung inflammation and fibrosis in diabetic rats, a synergistic effect of NAC and glycine in mitigating pulmonary complications associated with type 1 diabetes may be suggested. These findings warrant further exploration of the combination for managing diabetic lung disease and potentially other fibrotic conditions.

Amelioration of metabolic and behavioral defects through base editing in the PahR408W phenylketonuria mouse model

Phenylketonuria (PKU) is a liver metabolic disorders mainly caused by a deficiency of the hepatic phenylalanine hydroxylase (PAH) enzyme activity, often leading to severe brain function impairment in patients if untreated or if treatment is delayed. In this study, we utilized dual-AAV8 vectors to deliver a near PAM-less adenine base editor variant, known as ABE8e-SpRY, to treat the PahR408W PKU mouse model carrying a frequent R408W mutation in the Pah gene. Our findings revealed that a single intravenous injection in adult mice and a single intraperitoneal injection in neonatal mice resulted in 19.1% to 34.6% A-to-G editing efficiency at the pathogenic mutation site with minimal bystander edits. Furthermore, the dual-AAV8 treated mice exhibited reduced blood Phe levels to below the therapeutic threshold of 360 μmol L-1 and restored weight and fur color to normal levels. Importantly, the brain function of the mice was restored after the treatment, particularly when administered during the neonatal stage, as levels of monoamine neurotransmitters and metabolites in the brain returned to normal and near-normal levels. Our study demonstrated that ABE8e-SpRY-based base editing could effectively correct the point mutation in the PahR408W PKU mouse model, indicating potential clinical applications for PKU and other genetic diseases.

Isatin improves oligoasthenospermia caused by busulfan by regulating GSH/GPX4 axis to inhibit ferroptosis.

Ferroptosis, induced by iron overload and an imbalance in redox homeostasis, promotes the generation of reactive oxygen species (ROS), leading to iron-dependent lipid peroxides (LPO) and oxidative stress. Lipid peroxidation induced by reactive oxygen species is essential for the progression of spermatogenesis. However, its imbalance can lead to reproductive system damage and oligoasthenospermia, a critical cause of oligoasthenospermia. Isatin (ISA) is a naturally occurring compound that is widely distributed in lobsters, crustaceans, shellfish and various plants. It exhibits significant antioxidant and anti-aging properties, suggesting its potential as a therapeutic agent for the treatment of oligoasthenospermia. This study aimed to investigate the effects and mechanisms of ISA on oligoasthenospermia and to elucidate the underlying molecular pathways. All mice were divided into normal group, model group and treatment group. Both model group and treatment group received a single intraperitoneal injection of 30mg/kg BUS to create the model of oligoasthenospermia. After 2weeks, the treatment group received different doses of 25, 50 and 100mg/kg ISA by gavage for 28days, and then mice were sacrificed and tested. The results demonstrated that ISA effectively reversed busulfan-induced reproductive system damage in mice. This included the restoration of testicular histomorphology, improvement in sperm concentration and motility, regulation of serum sex hormone levels, and normalization of various oxidative indices in testicular tissue. Furthermore, ISA successfully reversed testicular ferroptosis by restraining the translocation of nuclear factor erythroid 2-related factor 2 (NRF2) into the nucleus and improved oligoasthenospermia through the glutathione (GSH)/glutathione peroxidase 4 (GPX4) axis. ISA was found to effectively ameliorate oligoasthenospermia in mice, presenting a potential therapeutic option for patients with this condition.

GLP-1 receptor agonist liraglutide alleviates kidney injury by regulating nuclear translocation of NRF2 in diabetic nephropathy.

Diabetic nephropathy (DN) is a severe renal disorder that arises as a complication of diabetes. Liraglutide, an analogue of a glucagon-like peptide 1 (GLP-1) receptor agonist, has been shown to decrease diabetes-caused renal damage. Nevertheless, the complete understanding of the roles and mechanism remains unclear. In our study, diabetic rat models were created through a single intraperitoneal injection of streptozotocin (STZ). The level of fasting blood glucose, 24-h urine protein, serum creatinine (Scr) and blood urea nitrogen (BUN) were assessed. Periodic acid-Schiff (PAS) staining was applied to examine the pathological changes in renal tissues. Reactive oxygen species (ROS) formation was measured via dichloro-dihydro-fluorescein diacetate (DCFH-DA) probes. Western blot was conducted to examine the levels of oxidative stress-related and extracellular matrix (ECM)-associated proteins. The nuclear translocation of NRF2 was investigated through immunofluorescence and Western blot assays. We demonstrated that liraglutide attenuated DN-induced oxidative stress and ECM deposition in vitro and in vivo. Liraglutide exerted a reno-protective effect by promoting nuclear translocation of NRF2 in mesangial cells. ML385, an NRF2 inhibitor, counteracted the beneficial impact of liraglutide.

Experimental study on the treatment of diabetic cardiomyopathy in rats by using ultrasound-targeted microbubble destruction combined with coenzyme Q10 loaded long-circulating nanoliposomes

Objective: To investigate the therapeutic effects and mechanisms of Ultrasound-targeted microbubble destruction (UTMD) technology combined with CoQ10 loaded PEGylated nanoliposomes (CoQ10-PEG-lips) on diabetic cardiomyopathy (DCM) in rats. Methods: CoQ10-PEG-lips were prepared using the thin-film dispersion method combined with ultrasonic hydration, followed by quality assessment. Sixty healthy and clean male SD rats were selected, and 50 were randomly chosen using a random number table to establish a type 1 diabetes mellitus (DM) model via a single intraperitoneal injection of streptozotocin. The remaining 10 rats were assigned as the normal control group. A total of 47 rats successfully developed the DM model, and 40 were selected using the random number table method. Based on different intervention methods, these rats were then randomly divided into DM model group, CoQ10 solution group, CoQ10-PEG-Lips group, and CoQ10-PEG-Lips+UTMD group (n=10 per group). Normal control group and DM model group rats were injected with 1 ml of normal saline through caudal vein. CoQ10 solution group and CoQ10-PEG-Lips group were injected with 1 ml of CoQ10 solution or CoQ10-PEG-Lips solution containing 10 mg/kg of CoQ10 through caudal vein, respectively. Rats in the CoQ10-PEG-Lips+UTMD group were injected with 1 ml of CoQ10-PEG-Lips solution containing 10 mg/kg of CoQ10+100 mg of freeze-dried ultrasound microbubble powder through caudal vein and were given UTMD treatment at the same time, with the intervention given twice a week. After 12 weeks of intervention, cardiac function indexes [left ventricular end-systolic diameter (LVEDs), left ventricular end-diastolic diameter (LVEDd), and left ventricular ejection fraction (LVEF)]of each group were measured by ultrasonic cardiac function detection in vivo. Simultaneously, ex-vivo histopathological examination was conducted to assess myocardial cell morphology, cross-sectional area, collagen volume fraction (CVF), and apoptosis index (AI) in each group. Additionally, molecular biology techniques were employed to measure oxidative stress-related indicators and the expression of apoptosis-related pathway proteins. Results: The prepared CoQ10-PEG-lips had a well-rounded morphology, good dispersibility, and a high encapsulation efficiency of 87.45%±3.23%. After 12 weeks of intervention, the myocardial cell morphology in the CoQ10-PEG-Lips+UTMD group was intact, with orderly arrangement, closely resembling that of the normal control group. There were no statistically significant differences between the two groups in terms of LVEDs [(1.53±0.07) mm vs (1.42±0.04) mm], LVEDd [(2.93±0.15) mm vs (2.81±0.05) mm], or LVEF (80.76%±3.42% vs 84.60%±2.10%) (all P>0.05). Similarly, there were no significant differences between the two groups in myocardial cell cross-sectional area, CVF, or AI (all P>0.05). The CoQ10-PEG-Lips+UTMD group showed statistically significant differences in the above-mentioned indicators compared to the DM group, CoQ10 solution group, and CoQ10-PEG-Lips group (all P<0.05). In the CoQ10-PEG-Lips+UTMD group, the levels of myocardial superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and the expression of the anti-apoptotic protein B-cell lymphoma-2 (Bcl-2) were significantly higher compared to the DM model group, CoQ10 solution group, and CoQ10-PEG-Lips group (all P<0.05), while malondialdehyde levels and the expression of Bcl-2-associated X protein (Bax) and caspase-3 were lower (all P<0.05). Conclusions: Using PEG-lips to encapsulate the poorly soluble drug CoQ10 in combination with UTMD technology enables targeted delivery of the drug to the myocardium, which can help reduce myocardial cell damage, fibrosis, and apoptosis caused by diabetes mellitus (DM) by inhibiting oxidative stress damage and the Bcl-2/Bax/caspase-3 apoptosis signaling pathway, ultimately improving cardiac function in DCM rats.