Male Sprague-Dawley Rats Research Articles

Ligustrazine nanoparticles inhibits epithelial-mesenchymal transition and alleviates postoperative abdominal adhesion

Following abdominal surgery, the occurrence of postoperative abdominal adhesion (PAA) is highly prevalent and stands out as one of the most frequently encountered complications. The effect and molecular mechanisms of Ligustrazine nanoparticles (LN) underlying epithelial-mesenchymal transition (EMT) in PAA still remain elusive.Adhesions were induced in Male Sprague-Dawley rats by injuring the cecum (cecal abrasion model), followed by administration of LN and hyaluronate acid (HA). The mechanism was further verified by enzyme-linked immunosorbent assay, wound healing assay, si-RNA and Western blot. Animal experiments revealed that LN effectively ameliorated adhesions, notably decreased tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), IL-8, and fibrosis, and reduced the expression of TGF-β1 and EMT related markers (Fibronectin and E-cadherin). Furthermore, in vitro experiments demonstrated that LN might inhibit the TGF-β1 FOXC2 pathway through suppressing the expression of Fibronectin, P120, and E-cadherin and ameliorating peritoneal adhesion. Collectively, our findings indicate that LN inhibits PAA formation by reducing inflammation, decreasing EMT and promoting peritoneal mesothelial cell repair. Therefore, LN might be considered a potential candidate for the treatment of PPA. However, further clinical studies are required to approve the effectiveness of LN.

The role of low-carbohydrate, high-fat diet in modulating autophagy and endoplasmic reticulum stress in aortic endothelial dysfunction of metabolic syndrome animal model.

Endothelial dysfunction (ED) is induced by insulin resistance, mediated by endoplasmic reticulum (ER) stress and disturbed autophagy. This study investigates the protective role of a low-carbohydrate, high-fat (LCHF) diet on ED, ER stress, and autophagy dysregulation in an experimental animal model of metabolic syndrome. Forty male Sprague-Dawley rats were divided into four groups: a Control group (standard diet) and three Dexamethasone (DEX) treated groups. Group II continued the standard diet, Group III received an LCHF diet, and Group IV received a high-carbohydrate, low-fat (HCLF) diet. At the end of the experiment, aortic tissue samples were obtained and used for histological, immunohistochemical (Endothelin and PCNA, biochemical MDA, TCA, NO, 8-OH-dG, and Nrf2/ARE protein) and molecular (Endothelin, eNOS, Nrf-2 α, p62, LC3, BECN-1, PINK1, CHOP, BNIP3, PCNA) analysis. Oxidative stress, autophagy markers, and ED markers are increased in the metabolic syndrome group. LCHF diet mitigates the adverse effects of DEX on endothelial dysfunction and oxidative stress, as evidenced by reduced BMI, HOMA-IR, and improved histological and molecular parameters. Oxidative stress, autophagy dysregulation, and ER stress play crucial roles in the pathogenesis of insulin resistance-induced endothelial dysfunction. An LCHF diet offers protective benefits against insulin resistance and related comorbidities, including endothelial dysfunction.

Therapeutic potential of resveratrol and autologous chondrocytes in male rat knee joint cartilage repair

Osteochondral defects (OCDs) in synovial joints are caused by trauma or inflammatory joint diseases, with no definitive treatment available. This study examined the effects of resveratrol and chondrocyte injections in a rat model of OCD. Twenty-four male Sprague-Dawley rats were divided into four groups: a control group, a resveratrol-only group (10 mg/kg), a chondrocyte-only group (1 × 105 cells), and a combined treatment group that received both treatments. After two months, the rats were euthanized, and their knee joints were analysed histologically and immunohistochemically. The results showed that the combined resveratrol and chondrocyte treatment significantly reduced fibrous tissue, increased cartilage tissue volume, improved cellular distribution, and enhanced the regularity of the articular surface. Collagen types I and II and proteoglycan levels were also elevated. These findings suggest that the combination of resveratrol and chondrocytes has a synergistic effect, promoting effective OCD repair in this rat model, offering potential for future therapeutic approaches.

Caffeic Acid Attenuates Neuronal Apoptosis, Oxidative Stress, and Memory Deficits via Antioxidant Properties in Aging Rats Induced by D-Galactose.

Aging is a main factor related to cognitive deficits. D-Galactose (D-gal), a monosaccharide, increases oxidative stress leading to cellular senescence, memory deficits, and neuronal apoptosis. Caffeic acid (CA) is an antioxidant that can interrupt free radicals and reduce oxidative stress. The present study purposely evaluated the benefits of CA in attenuating loss of neuronal apoptosis, oxidative stress, and memory in D-gal-activated rat brain aging. Male Sprague-Dawley rats were arbitrarily allocated into 6 groups (9 rats per group). The D-gal group was intraperitoneal (i.p.) injected with D-gal (50mg/kg). The CA groups were orally given 20 or 40mg/kg CA for 8weeks. During that time, the co-treatment groups were given 50mg/kg of D-gal and 20 or 40mg/kg of CA. The results reveal that animals receiving only D-gal showed memory deficit in both the novel object location (NOL) and novel object recognition (NOR) tests. Reduction in scavenging enzyme activities and levels of B-cell lymphoma 2 (Bcl-2) protein expression were detected in the D-gal group. Furthermore, D-gal treatment significantly enhanced in the number of p21 positive cells in the subgranular zone (SGZ) of the hippocampal dentate gyrus, Bcl-2 associated X protein (Bax) and caspase3 protein expression, and malondialdehyde (MDA) levels. By contrast, both 20 and 40mg/kg CA treatment alleviated these effects. These consequences confirmed that D-gal-activated brain aging led to enhancing apoptotic protein expression including Bcl-2, Bax, and caspase3 and memory impairments. Nevertheless, CA attenuated these effects in brain aging induced by D-gal via antioxidant properties.

Levamisole Ameliorates Rheumatoid Arthritis by Downregulating the PI3K/Akt Pathway in SD Rats.

Background/Objectives: Rheumatoid arthritis (RA) is a systemic chronic autoimmune disease characterized by a protracted course, high rates of morbidity, and disability yet lacks effective therapeutic modalities. Levamisole (LVM), an immunomodulatory drug, has been clinically reported for its potential in RA treatment, while its therapeutic mechanism toward RA remains to be elucidated. Hence, this study provides theoretical support for the application of LVM in the treatment of RA. Methods: This study employed male Sprague-Dawley (SD) rats to construct the adjuvant-induced arthritis (AIA) model, administering LVM orally (5 mg/kg, 15 mg/kg, and 45 mg/kg) for 25 days. An evaluation of LVM's therapeutic effects on RA was conducted through arthritis index scores, paw pad thickness, paw volume, hematoxylin and eosin (H&E) staining, 3D microcomputed tomography (micro-CT) scans, serum levels of pro-/anti-inflammatory cytokines, and serum biochemical indicators. Western blotting and immunohistochemistry staining were utilized to measure the expression levels of phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) proteins in synovial and ankle joint tissues. Results: Treatment with the median dose of LVM (15 mg/kg, M-LVM) significantly reduced the arthritis index (p < 0.01), paw pad thickness (p < 0.001), and paw volume (p < 0.01) without affecting body weight. Additionally, M-LVM alleviated inflammatory lesions in the synovium and ankle joints and also normalized serum levels of interleukin-1 beta (IL-1β), tumor necrosis factor-alpha (TNF-α), and transforming growth factor-beta (TGF-β). The Model group exhibited significant increases in serum levels of alkaline phosphatase (ALP) (p < 0.01), creatine kinase (CK) (p < 0.05), and glucose (GLU) (p < 0.001) compared with the Control group; however, M-LVM effectively regulated these parameters to normal levels. Western blotting and immunohistochemistry staining revealed that PI3K-/Akt-related proteins were highly expressed in the synovial and ankle joint tissues of rats in the Model group, while treatment with M-LVM significantly reduced the expression of these proteins. Furthermore, histological examination of major organs (heart, liver, lungs, kidneys, and thymus) showed no significant pathological changes, with the exception of the spleen, where M-LVM ameliorated splenic lesions. Conclusions: We demonstrate that LVM at an optimal dose substantially relieves synovitis and bone erosion in AIA rats by inhibiting the PI3K/Akt signaling pathway.

Sustained Protraction Increases Lengthening of the Mandibular Condylar Process Whilst Changes Its Growth Direction in a Rat Model.

In Class II elastics treatment, the mandible is subject to sustained protraction. The study aimed to investigate the morphological and histological changes resulting from sustained mandibular protraction, through establishment of a novel model in rat. Forty-eight 4-week-old male Sprague-Dawley rats were randomly divided into three groups, including the 50 and 100 cN protraction groups, and the control. In the two protraction groups, NiTi coil springs were fixed between the metal bands on the maxillary incisors and mandibular molars to pull the mandible forward. CBCT scans were taken immediately before (T0), 2 weeks (T1) and 4 weeks (T2) after the traction. Histological and immunohistochemical analyses were performed at T1 and T2. Increase of the condylar process length and angular process length were significantly larger in the two protraction groups than that in the control; however, the condylar process angle (∠CPA) was significantly reduced by the protraction at T2, leading to decreased condylar height. The protraction decreased the condylar cartilage thickness at T1, which recovered to the control level at T2, with no significant changes detected in the expression of SRY-related high mobility group-box gene 9 (SOX9) and type II collagen (COL II), two chondrogenic markers. The established rat model is manoeuvrable and reliable, which exerts sustained and measurable mandibular protraction. The sustained protraction increases the lengthening of the mandibular condylar process, but alters its growth direction by causing it to turn downwards.

Acute high-intensity muscle contraction moderates AChR gene expression independent of rapamycin-sensitive mTORC1 pathway in rat skeletal muscle.

The relationship between mechanistic target of rapamycin complex 1 (mTORC1) activation after resistance exercise and acetylcholine receptor (AChR) subunit gene expression remains largely unknown. Therefore, we aimed to investigate the effect of electrical stimulation-induced intense muscle contraction, which mimics acute resistance exercise, on the mRNA expression of AChR genes and the signalling pathways involved in neuromuscular junction (NMJ) maintenance, such as mTORC1 and muscle-specific kinase (MuSK). The gastrocnemius muscle of male adult Sprague-Dawley rats was isometrically exercised. Upon completion of muscle contraction, the rats were euthanized in the early (after 0, 1, 3, 6 or 24h) and late (after 48 or 72h) recovery phases and the gastrocnemius muscles were removed. Non-exercised control animals were euthanized in the basal state (control group). In the early recovery phase, Agrn gene expression increased whereas LRP4 decreased without any change in the protein and gene expression of AChR gene subunits. In the late recovery phase, Agrn, Musk, Chrnb1, Chrnd and Chrne gene expression were altered and agrin and MuSK protein expression increased. Moreover, mTORC1 and protein kinase B/Akt-histone deacetylase 4 (HDAC) were activated in the early phase but not in the late recovery phase. Furthermore, rapamycin, an inhibitor of mTORC1, did not disturb changes in AChR subunit gene expression after muscle contraction. However, rapamycin addition slightly increased AChR gene expression, while insulin did not impact it in rat L6 myotube. These results suggest that changes in the AChR subunits after muscle contraction are independent of the rapamycin-sensitive mTORC1 pathway.

Black Garlic Supplementation and Glomerular Protection in Hyperuricemic Rats: A Study on Kidney Health Prevention

Introduction: Hyperuricemia is a significant risk factor for the development of chronic kidney disease. Excessive uric acid can deposit in the kidneys, triggering inflammatory reactions and oxidative stress, leading to structural damage in the glomeruli. Black garlic, with its antioxidant and bioactive compounds, has the potential to protect the kidneys from hyperuricemia-induced damage. Our objective is to evaluate the effect of black garlic supplementation on the histopathological features of glomeruli in a hyperuricemic rat model. Methods: This study was an analytical observational study using Biological Remnant Materials (BRM) from a previous study. The sample consisted of 30 male Sprague-Dawley rats divided into 5 groups: disease control, drug control (allopurinol), and 3 black garlic dose groups (240 mg/day, 480 mg/day, and 960 mg/day). Data were collected through histopathological examination of kidney tissue using the glomerular damage scoring method (0-3). Data analysis was performed using one-way ANOVA and post hoc LSD tests. Results: Black garlic demonstrated protective effects on the histopathological features of glomeruli in hyperuricemic rats. The 240 mg/day dose showed the most significant improvement in kidney health, reducing glomerular damage scores compared to the disease control group (p &lt; 0.05). These findings suggest that black garlic can serve as a preventive agent against kidney complications caused by hyperuricemia. Conclusion: Black garlic provides protective effects on glomerular histopathology in hyperuricemic rats, with the 240 mg/day dose being the most effective. These results suggest potential use for black garlic in preventing kidney complications in hyperuricemia.

HES1 revitalizes the functionality of aged adipose-derived stem cells by inhibiting the transcription of STAT1.

The effectiveness of adipose-derived stem cells (ADSCs) in therapy diminishes with age. It has been reported that transcription factors (TFs) play a crucial role in the aging and functionality of stem cells. Nevertheless, there is limited understanding regarding the involvement of TFs in the aging mechanism of ADSCs. RNA sequencing (RNA-seq) was utilized to discern the differentially expressed genes in ADSCs obtained from donors of varying ages. TFs exhibiting significant variations across age groups were identified and subsequently validated. ADSCs were manipulated to exhibit either enhanced expression or reduced levels of HES1 and STAT1 via lentivirus transfection and small interfering RNA (siRNA) techniques. The impact of these genetic alterations on ADSCs' proliferation, migration, and cellular senescence was assessed using EdU, transwell, and senescence-activated β-galactosidase (SA-β-gal) staining assays. The DNA sequences bound by HES1 were investigated through the CUT & Tag assay. Lastly, the therapeutic efficacy of aged ADSCs with HES1 overexpression was evaluated in skin injury model of male Sprague-Dawley rats. 678 genes showed differential expression between ADSCs obtained from young and old donors (Y-ADSCs and O-ADSCs), with 47 of these genes being TFs. Notably, the expression of the TF hairy and enhancer of split 1 (HES1) was notably reduced in ADSCs from old donors. Introducing HES1 overexpression in aged ADSCs resulted in improved cellular function and the suppression of cellular senescence, while reducing HES1 levels in young ADSCs had the opposite effect. Mechanistically, HES1 was found to interact with the promoter region of another TF, signal transducer and activator of transcription 1 (STAT1), to inhibit its transcription. Knocking down STAT1 could fully reverse the negative effects caused by decreased HES1 in ADSCs, leading to a reduction in the secretion of pro-inflammatory cytokines such as TNF-α, IL-6, and IL-8. Ultimately, restoring HES1 expression in aged ADSCs demonstrated enhanced therapeutic potential in promoting skin wound healing. HES1 acts as an inhibitor of cellular senescence in the aging progression of ADSCs through the modulation of STAT1 expression, suggesting a promising avenue for rejuvenating senescent ADSCs and improving wound healing.

Post-exercise hot-water immersion is not effective for ribosome biogenesis in rat skeletal muscle.

Ribosome biogenesis is an important regulator of skeletal muscle hypertrophy induced by repeated bouts of resistance exercise (RE). Hot-water immersion (HWI), a widely used post-exercise recovery strategy, activates the mechanistic target of rapamycin (mTOR) signaling, a key regulator of ribosome biogenesis in skeletal muscle. However, the effect of HWI on skeletal muscle ribosome biogenesis is not well understood. Here, we aimed to investigate the effects of HWI and post-exercise HWI on ribosome biogenesis using a rat RE model. Male Sprague-Dawley rats were randomly assigned to HWI and non-HWI groups. In both groups, the right leg was isometrically exercised using transcutaneous electrical stimulation, while the left leg was used as an internal non-RE control. Following RE, both limbs were immersed in hot water (41.2 ± 0.03℃) for 20 min under isoflurane anesthesia in the HWI group and the gastrocnemius muscles were sampled at 3 and 24 h post-exercise. HWI significantly increased mTOR signaling and c-Myc mRNA expression, whereas post-exercise HWI significantly increased transcription initiation factor-IA mRNA expression. However, neither HWI nor post-exercise HWI enhanced 45S pre-rRNA expression, ribosomal RNA, or ribosomal protein content. Additionally, HWI tended to decrease 28S rRNA and 18S rRNA content, widely used markers of ribosome content. These results suggest that HWI as a post-exercise recovery is not effective in activating ribosome biogenesis.

Endoplasmic Reticular Stress and Pathogenesis of Experimental Colitis: Mechanism of Action of 5-Amino Salicylic Acid.

Inflammatory bowel diseases which are characterized by endoplasmic reticulum (ER) stress and activation of the unfolded protein response (UPR) signaling pathway, are commonly treated with 5-amino salicylic acid (5-ASA). The objective of this study was to investigate the role of 5-amino salicylic acid in the UPR-signaling pathway in experimental colitis. Colitis was induced in male Sprague-Dawley rats by intrarectal instillation of trinitrobenzene sulfonic acid. Animals received 5-amino salicylic acid (100 mg/kg body weight) two hours before the induction of colitis and repeated daily until day 7. The animals were sacrificed on day 7 and tissues were collected for analysis. The expression of protein kinase R (PKR)-like endoplasmic reticulum (ER) kinase (PERK), a mediator of UPR-signaling increased significantly (p < 0.05), while inositol requiring enzyme type-1 (IRE1) and the CCAAT/enhancer-binding homologous protein (CHOP) remained unaltered in the inflamed colon. The expression of glucose regulated protein-78, activator of transcription factor-4 and phosphorylated eukaryotic initiation factor-2α (eIF2αP) increased (p < 0.05) in the inflamed colon. However, the levels of eIF2α protein and mRNA expression remained unchanged. Myeloperoxidase activity, colon weight and infiltration of inflammatory cells increased significantly (p < 0.05) in the submucosa whereas the body weight decreased. These changes were significantly inhibited by 5-amino salicylate treatment. These findings suggest that the anti-inflammatory properties of 5-amino salicylic acid are mediated through the inhibition of the PERK signaling pathway.

Asiatic acid alleviates subarachnoid hemorrhage-induced brain injury in rats by inhibiting ferroptosis of neurons via targeting acyl-coenzyme a oxidase 1

The occurrence of subarachnoid hemorrhage (SAH) can lead to brain injury, which is a fatal condition with limited effective clinical intervention strategies. The naturally occurring component Asiatic acid (AA), found in the tropical plant Centella asiatica, has been reported to possess neuroprotective properties. The objective of this study was to evaluate the neuroprotective effect of AA following SAH and investigate its potential mechanisms. The SAH model was established in male Sprague-Dawley (SD) rats through intravascular perforation, following a standardized protocol. The administration of AA was performed via gavage following SAH. A lentiviral vector was constructed and utilized for the knockdown of Acyl Coenzyme A Oxidase 1 (ACOX1) Firstly, AA treatment effectively improves brain neurological deficit, neuronal damage, and iron deposition induced by SAH. Furthermore, it has been demonstrated that AA directly interacts with ACOX1, which exhibits decreased expression in neurons following SAH. Additionally, our study reveals AA can reverse SAH-induced reduction in ACOX1 expression, concurrently ameliorating neuronal ferroptosis. This improvement is evidenced by reduced lipid peroxidation, including mitigated GSH depletion, decreased MDA production, and increased GPX4 content and activity. Also, AA enhances mitochondrial constriction while alleviating cristae disruption induced by SAH, providing crucial insights into its neuroprotective effects against neuronal ferroptosis in SAH. Moreover, when ACOX1 is knocked down, the neuroprotective effects of AA are weakened. Collectively, this study elucidated the neuroprotective effect of AA by inhibiting neuronal cell ferroptosis through targeting ACOX1. These findings suggest that AA holds promise as a potential therapeutic candidate for ameliorating SAH-induced brain injury.

Integrated gut microbiome and UHPLC-MS metabolomics to reveal the prevention mechanism of pidanjiangtang granules on IGT Rats

IntroductionPidanjiangtang (PDJT) is a traditional Chinese medicine formula empirically used to treat impaired glucose tolerance (IGT) based on the "Pidan" theory from the classic ancient book Nei Jing. However, the mechanism of PDJT intervention for IGT remains to be studied. ObjectiveThis study aims to explore the mechanism of PDJT granules intervention in IGT by integrating gut microbiome and UHPLC-MS untargeted metabolomics. Materials and methodsThe IGT model was established in 6-week-old male Sprague-Dawley (SD) rats by feeding them a high-fat diet and using an STZ injection. The low, medium, and high doses of PDJT were used for six weeks. metformin (Glucophage) was used as the positive control drug. The efficacy of PDJT was evaluated using fasting blood glucose (FBG), blood glucose maximum (BGmax), blood lipid, and inflammatory factor levels. Finally, 16S rDNA gut microbiome sequencing with metabolomics analysis was used to explore the pharmacological mechanism of PDJT intervention in IGT. ResultsPDJT could reverse the phenotype of IGT rats, reduce blood glucose levels, improve lipid metabolism disorder, and reduce inflammatory response. Gut microbiome analysis found that PDJT can improve gut microbiota composition and abundance of three phyla (Firmicutes, Bacteroidota, Desulfobacterota) and four genera (unclassified_f__Lachnospiraceae, Ruminococcus, Allobaculum, Desulfovibrio), which play an important role in the process of PDJT intervention on glucose metabolism and lipid metabolism in IGT rats. UHPLC-MS untargeted metabolomics showed that PDJT could regulate the levels of 258 metabolites in lipid metabolism pathways, inflammatory response pathways, fat and protein digestion, and absorption. The combined analysis of the two omics showed that improving the body's metabolism by gut microbes may be the possible mechanism of PDJT in treating IGT. Thus, this study provides a new method to integrate gut microbiome and UHPLC-MS untargeted metabolomics to evaluate the pharmacodynamics and mechanism of PDJT intervention in IGT, providing valuable ideas and insights for future research on the treatment of IGT with traditional Chinese medicine.

Investigating Anti-Oxidative and Anti-Inflammatory Effects regarding Ethanol Extract of Allium Porrum L. in Rats with Type 2 Diabetes Mellitus

Background: Type 2 diabetes mellitus (T2D) is a common endocrine and metabolic disorder worldwide. Meanwhile, Allium Porrum L. (AP) has been recognized as one of the most eminent traditional herbal medicines with substantial health benefits. The aim of the present study is to evaluate anti-oxidative and anti-inflammatory effects of ethanol extract of AP in rats with T2D. Methods: 70 male Sprague-Dawley rats were randomly divided into 7 groups of 10 animals each. The study groups were as follows: The study groups were as follows: healthy control (HC); healthy control rats receiving 100 mg/kg of AP extract (HC.AP.100); diabetic control (DC); diabetic rats receiving 10 mg/kg metformin (DT.M.10); diabetic animals receiving 50 mg/kg of AP extract (DT.AP.50); diabetic rats receiving 100 mg/kg of AP extract (DT.AP.100); and diabetic animals receiving 200 mg/kg of AP extract (DT.AP.200). Results: After 6 weeks of intervention, the level of serum malondialdehyde (MDA) and tumor necrosis factor alpha (TNF-α) were significantly increased (P=0.02) and (P=0.002), respectively; while superoxide dismutase (SOD) concentration was notably reduced (P=0.04) in DC.M.10 in comparison with that of the HC.AP.100 group. Also, compared to DC.M.10 group, all rats treated with metformin and AP showed a significant decrease in the level of MDA and TNF-α, along with an enhancement of SOD concentration (All P&lt;0.001). Moreover, in comparison with DT.M.10, DT.AP.50, and DT.AP.100 treatments, the DT.AP.200 group revealed a significantly higher improvement in serum TNF-α concentration (P=0.04, P=0.003 and P=0.003, respectively). Conclusion: The findings revealed that the use of AP extract for 6 weeks may have beneficial effects on oxidative stress and inflammatory biomarkers in T2D-induced rat models.

Fecal microbiota transplantation alleviates neuronal Apoptosis, necroptosis and reactive microglia activation after ischemic stroke

ObjectiveThis study aims to delve into the mechanisms underlying the improvement of neurological function in rats with ischemic stroke through fecal microbiota transplantation. MethodsA total of fifty male Sprague-Dawley rats were categorized into four groups: Sham, MCAO, MCAO+vehicle and FMT. We assessed behavioral and pathological alterations in the rats using modified neurological function scoring and TTC staining.Additionally, Western blot and immunofluorescence were used to detect the expression levels of Apoptotic and Necroptosis markers in neurons of ischemic brain tissue, and immunofluorescence was used to analyze the degree of activation of microglia. ResultsFMT group exhibited a decline in neurological function score compared to the MCAO and MCAO + vehicle group, accompanied by a reduction in infarct volume (P < 0.05). Relative to the SHAM group, the MCAO group displayed a significant increase in the expression levels of necroptosis-related proteins Phospho-RIP1, Phospho-RIP3, Phospho-MLKL, apoptotic proteins Bax and Cleaved caspase-3, and the iNOS positive microglia in ischemic brain tissue, while Bcl-2 expression was notably decreased (P < 0.05).Conversely, compared to the MCAO + vehicle group, the FMT group showed decreased expression levels of Phospho-RIP1, Phospho-RIP3, Phospho-MLKL, Bax, Cleaved caspase-3, and iNOS-positive microglia, while the expression of Bcl-2 was increased. ConclusionFecal microbiota transplantation offers a promising approach to improving neurological function in rats with ischemic stroke by inhibiting neuronal apoptosis, necroptosis, and the polarization of inflammatory microglial cells.

Second generation multiple channeling using platelet-rich plasma enhances cartilage repair through recruitment of endogenous MSCs in bone marrow.

In the treatment of cartilage defects, a key factor is the adequate and specific recruitment of endogenous stem cells to the site of injury. However, the limited quantity and capability of endogenous bone marrow stem cells (BM MSCs) often result in the formation of fibrocartilage when using bone marrow stimulation (BMS) procedures. We engineered second-generation platelet-rich plasma (2G PRP) with defibrinogenating and antifibrinolytic agents for injection into the condyle of the right femur, followed by multiple channeling (MCh) 5 days later. This approach aims to enhance repair by promoting the local proliferation and migration of BM MSCs to the full-thickness knee cartilage defect (ftKD). In our in vitro study, 2G PRP increased the number of endogenous BM MSCs and their ability to migrate toward an IL-1β-induced inflammatory condition. This significance was further confirmed by in vivo proliferation results after injection of 2G PRP into the condyle of rats. Fifty-four healthy male Sprague-Dawley rats were divided into 3 groups (ftKD, MCh, 2G MCh) for 3 time points (2 weeks, 4 weeks, 8 weeks). The 2G MCh (2G PRP injection + MCh) groups significantly improved cartilage formation at 4 and 8 weeks compared to the ftKD and MCh groups. The 2G MCh initiated cartilage repair earlier than MCh and significantly enhanced up to 8 weeks. This study demonstrated that 2G PRP increased the number of BM MSCs through the enhancement of proliferation and recruitment into the injured site, thereby improving articular cartilage repair.

Efficacy of Low-Dose Ketamine and Propofol in the Treatment of Experimental Refractory Status Epilepticus on Male Rats.

Refractory status epilepticus (RSE) is a condition with serious mortality and morbidity rate, resistant to benzodiazepine and second-line antiepileptic drugs. This study aimed to electrophysiologically investigate the combination of NMDA receptor antagonist ketamine and GABAergic agent propofol in an RSE model induced by lithium-pilocarpine in male Sprague-Dawley rats. Seventy-two male Sprague-Dawley rats were divided into nine groups. The RSE model was induced by subcutaneous injection of lithium-CI (5 mEq/kg) and intraperitoneal injection of pilocarpine-HCl (320 mg/kg), after implanting tripolar EEG electrode. Ketamine (30, 60, and 90 mg/kg), propofol (20, 40, and 80 mg/kg), and combinations of both drugs (15 + 20 and 30 + 40 mg/kg) were administered intraperitoneally to animals with RSE. Video-EEG recordings were taken after inducing model and 48 h later. The efficacy of drugs was statistically evaluated based on spike frequencies (spikes/min) and amplitudes (mV). Compared to RSE group, it was determined that 30 and 60 mg/kg doses of ketamine provided effective seizure control and prevented mortality (p < 0.001), while the 90 mg/kg showed toxic effects in all animals and caused mortality. The 80 mg/kg dose of propofol provided seizure control and reduced the mortality rate to 16.7% (p < 0.001), whereas the 20 mg/kg resulted in a 100% mortality rate. The low-dose ketamine+propofol (15 + 20 mg/kg) combination provided early onset seizure control and were as effective as 80 mg/kg propofol (p < 0.05). The study concluded that in the experimental RSE model, seizure control could be achieved with low-dose combination of ketamine and propofol without the need for high doses as in monotherapy, thus preventing dose-related adverse effects.

The trans-differentiation promotion of parietal epithelial cells by magnesium isoglycyrrhizinate to improve podocyte injury induced by high fructose consumption

BackgroundPodocytes have limited proliferative capacity, which leads to irreversible glomerular injury in diverse kidney diseases. Magnesium isoglycyrrhizinate (MgIG), a hepatoprotective agent in clinic, has been reported to improve glomerular podocyte injury. However, the underlying mechanism of MgIG in ameliorating podocyte injury remains unclear. PurposeGlomerular parietal epithelial cells (PECs) are recognized as podocyte progenitors and play a pivotal role in the recovery following glomerular injury. This work aims to investigate the protective mechanisms of MgIG in mitigating glomerular injury by promoting PEC trans-differentiation. Study DesignA rat model of progressive glomerular podocyte injury, and in vitro models using the primary podocytes and primary PECs, were established to further explore the pharmacological mechanism of MgIG. MethodsFour-week-old male Sprague-Dawley (SD) rats were fed a 10% fructose solution for 3, 6, 9 and 12 weeks to induce glomerular injury. The effect of MgIG on the progressive changes in podocytes and PECs, and the correlation between PEC density and podocyte loss, were analyzed. The mechanism of MgIG in triggering PEC trans-differentiation was investigated, by examining adenosine secretion in injured podocytes, as well as the expression of cluster of differentiation 44 (CD44), nephrin, adenosine receptor A2B (ARA2B) and glucocorticoid receptor (GR) in PECs were examined both in vivo and in vitro. ResultsRats fed a high fructose diet exhibited progressive changes in glomerular PECs, including increased cell density and a preference for trans-differentiation. A positive correlation was observed between PEC density and podocyte loss. Co-culture experiments demonstrated that extracellular adenosine accumulation in injured podocytes induced by high fructose exposure promoted PEC trans-differentiation via ARA2B. MgIG significantly improved podocyte injury and exhibited effects similar to dexamethasone on nephrin upregulation activation and CD44 inhibition. Moreover, the inhibitory effect of MgIG on PEC ARA2B activation was more effective than that of dexamethasone. The co-expression of paired box 2 (PAX2)+-Nephrin+ in glomeruli indicated that MgIG induced PEC trans-differentiation and podocyte regeneration in model rats. Accordingly, podocyte loss and increased urine albumin-to-creatinine ratio (UACR) were also alleviated. Moreover, MgIG, which acts as a GR agonist to activate GR, reversed the upregulation of CD44 and decreased ARA2B induced by tumor necrosis factor-α (TNF-α) in primary PECs. The siRNA interference experiment manifested that MgIG exhibited a more pronounced enhancement of GR upregulation, in contrast to ARA2B activation, to promote PEC trans-differentiation. ConclusionThis work reports for the first time that PECs respond to the accumulation of extracellular adenosine from injured podocytes via activating ARA2B and focuses on the role of adenosine and adenosine receptors in the trans-differentiation of PECs. Furthermore, this study provides the first evidence that MgIG may promote podocyte regeneration by enhancing PEC trans-differentiation through GR activation, providing a research basis for investigating the glucocorticoid-like activity of MgIG in ameliorating glomerular podocyte injury.

Electroacupuncture Protects Against Cerebral Ischemia-Reperfusion Injury: Reducing Inflammatory Response and Cell Pyroptosis by Inhibiting P2X7/NLRP3/GSDMD Pathway.

To investigate the mechanism underlying the effects of Tongdu Tiaoshen electroacupuncture in the treatment of cerebral ischemia-reperfusion (I/R) injury. Sixty adult male Sprague-Dawley (SD) rats were randomly allocated to five groups (n=12): Sham, I/R, electroacupuncture (EA), BBG (P2X7R inhibitor), and MCC950 (NLRP3 inhibitor). The EA group received acupuncture at Shenting (GV24), Baihui (GV20), and Dazhui (GV14) points with a stimulation frequency of 2/5 Hz, intensity of 2 mA, and a duration of 40 min. Neurologic deficit scoring was then performed, and cerebral infarction volume was determined using triphenyl tetrazolium chloride (TTC) staining. Morphological changes in neurons in cortical areas were observed using an electron microscope, interleukin-18 (IL-18) and interleukin-1β (IL-1β) levels were assessed via the enzyme-linked immunosorbent assay (ELISA). Moreover, the expression of P2X7R/NLRP3/GSDMD pathway-related proteins and mRNAs was quantified by immunofluorescence staining, Western blot assay, and real-time fluorescence quantitative (RT-PCR) analysis. Electroacupuncture improved neurologic deficit scores, cerebral infarct size, and cell pyroptosis, and reduced IL-1β and IL-18 levels. Furthermore, electroacupuncture and inhibitor treatment significantly downregulated the expression of P2X7R, ASC, Caspase-1, NLRP3, and GSDMD involved in the P2X7/NLRP3/GSDMD signaling pathway. Tongdu Tiaoshen electroacupuncture may exert cerebral protection by inhibiting neuroinflammation through the P2X7/NLRP3/GSDMD pathway and reducing cellular pyroptosis.

Effect and related mechanism of acetate in alleviating acute kidney injury in septic rats through G-protein coupled receptor 43

To explore the protective effect and mechanism of acetate on sepsis-induced acute kidney injury (AKI) in rats. Male Sprague-Dawley (SD) rats were divided into sham operation group (Sham group), sepsis group caused by cecal ligation and puncture (CLP group), and acetate pretreatment group [NaA group, gavage sodium acetate (NaA) 300 mg/kg twice a day for 7 consecutive days before CLP] using a random number table method, with 7 rats in each group. The blood was taken from the main abdominal artery 24 hours after modeling, and renal tissue was collected from the rats. Enzyme linked immunosorbent assay (ELISA) was used to detect the serum levels of interleukin-6 (IL-6), tumor necrosis factor-α(TNF-α), and kidney injury molecule-1 (KIM-1). The concentration of serum acetate was determined by high performance liquid chromatography. The level of malondialdehyde (MDA) in renal tissue was detected by thiobarbituric acid method. Myeloperoxidase (MPO) in renal tissue was detected by colorimetric method. Hematoxylin-eosin (HE) staining was used to observe histopathological changes and assess renal tubule injury score. Western blotting was used to detect the protein expressions of G-protein coupled receptor 43 (GPR43) and adenosine monophosphate-activated protein kinase/silence infor-mation regulator 1/peroxlsome proliferator-activated receptor-γ coactlvator-1α (AMPK/SIRT1/PGC-1α) pathway. The positive expressions of GPR43, phosphorylation-AMPK (p-AMPK), SIRT1, PGC-1α were detected by immunohistochemistry. Compared with Sham group, the serum levels of IL-6, TNF-α and KIM-1 were significantly increased in CLP group, the contents of MDA and MPO in renal tissue were increased, and the content of acetate was significantly decreased. HE staining results showed that most of the tubular epithelial cells were denaturated with local necrosis, a large number of brush border injuries and shedding, tubular structure destruction and fragmentation, and more inflammatory cells infiltrated the renal interstitium, the renal tubular injury score significantly increased. The expressions of GPR43, p-AMPK/AMPK, SIRT1, and PGC-1α in renal tissue were significantly reduced, indicating renal injury and increased levels of oxidative stress and inflammation in septic rats. Compared with the CLP group, the serum levels of IL-6, TNF-α and KIM-1 in the NaA group were decreased [IL-6 (ng/L): 126.20±6.23 vs. 161.00±17.37, TNF-α (ng/L): 85.59±7.70 vs. 123.50±17.78, KIM-1 (μg/L): 2.92±0.38 vs. 4.73±0.36, all P < 0.05]. The contents of MDA and MPO in renal tissue were significantly decreased [MDA (μmol/g): 6.56±0.18 vs. 8.53±0.34, MPO (U/g): 2.99±0.20 vs. 3.72±0.29, both P < 0.05]. HE staining showed that kidney injury had been alleviated, with a decrease in renal tubular injury score [1 (1, 2) vs. 3 (2, 3), P < 0.05]. Western blotting showed that the expressions of GPR43 and AMPK/SIRT1/PGC-1α pathway related proteins were significantly increased in renal tissue (GPR43/β-actin: 0.62±0.09 vs. 0.41±0.09, p-AMPK/AMPK: 0.58±0.07 vs. 0.44±0.06, SIRT1/β-actin: 0.85±0.06 vs. 0.73±0.03, PGC-1α/β-actin: 0.79±0.07 vs. 0.62±0.05, all P < 0.05). Immunohistochemistry showed that the positive expressions of GPR43, p-AMPK, SIRT1 and PGC-1α were significantly increased in renal tissue [GPR43 positive area: (33.66±2.62)% vs. (16.21±1.66)%, p-AMPK positive area: (16.64±2.11)% vs. (5.04±1.28)%, SIRT1 positive area: (14.61±2.86)% vs. (7.34±1.00)%, PGC-1α positive area: (15.30±2.39)% vs. (4.84±1.67)%, all P < 0.05], the serum acetate concentration significantly increased (μg/L: 32 479±14 683 vs. 12 935±3 197, P < 0.05). Acetate can ameliorate sepsis-induced AKI, the mechanism may be related to the activation of AMPK/SIRT1/PGC-1α pathway by GPR43.