Periodic acid-Schiff Research Articles

Effects of Benzo[α]pyrene on Mucus Secretion and Tissue Remodeling in a Rat Model of Allergic Rhinitis.

Exposure to benzo[α]pyrene (BaP) increases the incidence and severity of allergic rhinitis (AR), but the underlying mechanisms remain unclear. Thus, we investigated the in vivo effects of BaP exposure on mucus hypersecretion and tissue remodeling in a rat model of AR. Female Sprague-Dawley rats were randomly divided into 4 groups: a negative control group, a group of healthy rats exposed to BaP, a group of rats with ovalbumin (OVA)-induced AR, and a group of AR model rats exposed to BaP. Nasal symptoms and levels of OVA-specific serum immunoglobulin E (IgE) were measured in each individual rat. Moreover, examination of goblet cell hyperplasia and collagen deposition was carried out with periodic acid-Schiff (PAS) staining and Masson trichrome (MT) staining. Mucin 5AC (MUC5AC) expression was assessed by immunohistochemistry. BaP significantly increased the number of sneezes, the number of nasal rubs and the levels of OVA-specific serum IgE in rats with AR. Statistically significant differences in goblet cell hyperplasia and collagen deposition were observed between the BaP-exposed AR model group and the AR model group. Immunohistochemical results showed that the nasal mucosa of AR model rats displayed markedly elevated MUC5AC expression after BaP exposure. Our data indicate that mucus hypersecretion and the development of nasal remodeling might be pathophysiologic mechanisms underlying increased susceptibility to AR after exposure to BaP.

Incidence and characterization of polyglucosan bodies in the cerebella of montserrat orioles (Icterus oberi).

Polyglucosan bodies are accumulations of insoluble glucose polymers and proteins that form intracytoplasmic inclusions in the brain, large numbers of which can be indicative of neurodegenerative diseases such as Lafora disease. Montserrat orioles (Icterus oberi) are an icterid passerine endemic to Montserrat with conservation populations maintained in captivity abroad. We demonstrate that polyglucosan bodies are unusually abundant in the cerebellar molecular and Purkinje cell layers and cerebellar peduncles of captive-bred and wild-caught Montserrat orioles. The bodies are periodic acid-Schiff positive and diastase resistant and label with concanavalin A and for ubiquitin, consistent with those seen in humans. We found no association of the polyglucosan bodies with concurrent neurological lesions or clinical signs, nor with EPM2A and EPM2B gene mutations associated with Lafora disease. We conclude that an abundance of cerebellar polyglucosan bodies may be a normal finding in aged Montserrat orioles and not a threat to the captive breeding population.

A Rare Case of Late Onset-Pompe’s Disease: Presented as Heart Failure

Pompe disease is a hereditary lysosomal storage disorder characterized by a deficiency in the acid alpha-glucosidase (GAA) enzyme, leading to glycogen accumulation in muscle and neurons. Clinical manifestations vary from severe infantile-onset with hypertrophic cardiomyopathy and early mortality due to respiratory insufficiency to late-onset with proximal muscle weakness, gross motor delay, and progressive respiratory insufficiency. A case of an 11-year-old boy who reported to the pediatric emergency department with a nine-year history of progressive muscle weakness and a one-month history of anemia symptoms (easy fatigue, shortness of breath, pale appearance) and heart failure (orthopnea, dyspnea). His family history included consanguineous marriages and similar conditions in his brother and maternal uncle. On examination, he appeared pale, malnourished, and exhibited signs of respiratory distress and tachypnea. His cardiovascular examination revealed apex beat displacement, elevated JVP, bilateral pedal edema, mild ascites, positive hepatojugular reflux, and systolic murmurs. Respiratory examination indicated bilateral crepitation and wheezes. Musculoskeletal examination showed decreased muscle mass and power, especially in proximal muscles. Abdominal examination revealed hepatosplenomegaly and mild ascites. Radiological findings included an enlarged cardiac shadow with pleural effusion and bilateral radio-opaque shadows on chest x-ray, while echocardiography showed impaired left ventricular systolic function with mild to moderate mitral and tricuspid regurgitation. Laboratory tests indicated elevated aspartate aminotransferase, LDH, and creatine kinase levels, along with normocytic, normochromic anemia. Muscle biopsy from the hamstring revealed PAS stain positive granules. These clinical, radiological, and laboratory findings strongly suggest late-onset Pompe disease, marking this as potentially the second reported case in Pakistan.

Fufang Zhenzhu Tiaozhi (FTZ) capsule ameliorates diabetic kidney disease in mice via inhibiting the SGLT2/glycolysis pathway

Ethnopharmacological relevanceFufang Zhenzhu Tiaozhi (FTZ) capsule is a hospital preparation of a patented traditional Chinese medicine compound. FTZ has been clinically used for nearly 13 years in the treatment of diabetes and glycolipid metabolic diseases. With the significant benefits of SGLT2 inhibitor in patients with diabetic kidney disease (DKD), it provides a research avenue to explore the mechanism of FTZ in treating this disease based on glycolysis pathway. Aim of the studyTo explore the pharmacological characteristics of FTZ in DKD mice and its impact on the glycolysis pathway. Materials and methodsWe induced a DKD model in C57BL/6 mice by injection of streptozotocin (STZ) combined with long-term high-fat diet. We administered three doses of FTZ for 12 weeks of treatment. Kidney function, blood lipid levels, glucose tolerance, and key glycolytic enzymes were evaluated. Renal pathological changes were observed using HE, MASSON, and PAS staining. The potential targets of the active ingredients of FTZ in the glycolysis pathway were predicted using network pharmacology and molecular docking. Validation was performed using immunohistochemistry and Western blotting. ResultsFTZ effectively reduces blood glucose, total cholesterol, triglyceride, low density lipoprotein cholesterol, 24 h proteinuria, serum creatinine, blood urea nitrogen, and increases urinary glucose levels. Glucose tolerance and renal pathological changes were significantly improved by FTZ treatment. Pinusolidic acid, a component of FTZ, shows good binding affinity with three active pockets of SGLT2. WB and immunohistochemistry revealed that FTZ significantly inhibits the expression of SGLT2 and its glycolytic related proteins (GLUT2/PKM2/HK2). Hexokinase, pyruvate kinase, and lactate dehydrogenase in the kidney were also significantly inhibited by FTZ in a dose-dependent manner. ConclusionFTZ may alleviate the progression of DKD by inhibiting the activation of the SGLT2/glycolytic pathway. Our study provides new insights into the clinical application of FTZ in DKD.

A new animal model of cardiorenal syndrome could be established by inducing heart failure through coronary artery ligation in spontaneously hypertensive rats

In rats with unilateral nephrectomy and cardiac dysfunction, renal function deteriorates at an accelerated rate, as evidenced by increased proteinuria. Whether myocardial infarct-induced heart failure (HF) exacerbates renal injury in hypertensive rats with mild renal injury has not been reported. Rats underwent either coronary ligation or sham surgery. Thirty spontaneously hypertensive rats (SHRs) aged 8 weeks were randomly divided into two groups. Group 1 was the sham group, in which the rats underwent thoracotomy without ligation of the coronary artery. Group 2 underwent coronary artery ligation. The rats in group 2 underwent coronary artery ligation on week 0. The experiment lasted 12 weeks. Urine was collected in metabolic cages over a 24-h period. Urine was collected from the rats 2 days before the end of the experiment, and the ratio of urinary protein to urinary creatinine was measured in the clinical laboratory. All rats were examined by echocardiogram one day before the end of the experiment. On the last day of the experiment, blood was collected and sent to the laboratory for analysis. Hematoxylin–eosin (HE) and periodic acid-Schiff (PAS) staining were performed on heart and kidney sections. The ejection fraction in group 2 was lower than that in group 1 (P < 0.001). The urinary albumin to creatinine ratio in group 2 was greater than that in group 1 (P < 0.001). The urea and creatinine levels in group 1 were significantly lower than those in group 2 (P < 0.01). The levels of brain natriuretic peptide (BNP), neutrophil gelatinase-associated lipocalin (NGAL) and cystatin C were greater in the second group than in the first group (P < 0.05). The interleukin-1β (IL-1β) and interleukin-6 (IL-6) levels in group 2 were significantly greater than those in group 1 (P < 0.001). The malondialdehyde (MDA) levels in Group 2 were greater than those in Group 1 (P < 0.01). The glutathione peroxidase (GSH-Px) levels in Group 2 were lower than those in Group 1 (P < 0.05). The level of angiotensin II (AT-II) in group 1 was lower than that in group 2 (P < 0.001). Cardiac dysfunction secondary to myocardial infarction could induce cardiorenal interactions in SHRs. It could be interpreted by the activation of oxidative stress, changes in inflammation and alteration of renin–angiotensin–aldosterone system.

Effect of fish oil supplementation on histological changes, apoptosis and oxidative stress of rat placenta against formaldehyde-induced toxicity

Formaldehyde (FA) as a common organic compound has been shown to cause placental dysfunction and fetal defects. The potential benefits of fish oil (FOil) in protecting placental structures are attributed to its antioxidant properties. This study aimed to explore the preventive role of FOil in mitigating the adverse effects of FA in pregnant rats. Thirty pregnant Wistar rats were randomly categorized into five groups of control, sham (Normal saline; Orally and intraperitoneally), FOil (0.5 ml/day; Orally), FA (5 mg/kg/bw; intraperitoneally), FA+FOil. The treatment period was from day 0–20 of pregnancy. On the 20th day of pregnancy, placental morphometric parameters were measured. The histological and histochemical analyses were performed using H&E and PAS staining, respectively. Also, the placenta tissue was analyzed for oxidative stress biomarkers, p-53 protein levels, and the expression of caspase-3 gene. The administration of FA led to a significant decrease in the weight, diameter, and thickness of the placenta, as well as a decrease in the thickness of the decidua layer, junctional and labyrinth zone, and the number of trophoblast giant cells in rat placentas. FA led to a significant increase in placental p-53 protein levels, caspase-3 expression, and oxidative stress biomarkers. Administration of FOil to pregnant rats treated with FA led to a significant decrease in morphometric and histological changes, oxidative stress, and the expression of genes associated with apoptosis. The findings suggest that the administration of FOil to FA-treated pregnant rats can protect placental histopathological changes by enhancing the activity of the antioxidant enzymes.

Potential of Bauhinia variegata leaves in the management of diabetic nephropathy

The present study was carried out to explore the potential of aqueous extract of leaves of Bauhinia variegata (AE) in the treatment of streptozotocin-induced diabetic nephropathy in rats. Diabetes was induced in male Sprague Dawley rats with streptozotocin (STZ) at a dose of 55 mg/kg (i.p.). Four weeks after induction of diabetes animals were treated with the extract at a dose of 250, 500 and 1000 mg/kg for 28 days. On the 28th day, biochemical parameters (glucose, glycosylated haemoglobin, total protein, albumin, creatinine, blood urea nitrogen) and urine parameters (total protein, creatinine) were evaluated. After completion of 28 days of treatment, animals were sacrificed and kidneys were isolated. Histopathological evaluation of kidney tissue was carried out by staining the tissue sections with hematoxylin and eosin (HE) stain, Periodic acid–Schiff (PAS) stain and Trichrome stain. Treatment with AE to diabetic rats for 28 days showed a significant decrease in fasting blood glucose, glycosylated haemoglobin, and biochemical parameter like albumin, total protein, creatinine, and blood urea nitrogen (BUN) which is an important marker for the progression of diabetic nephropathy. AE treatment also improved the urine parameters and histological changes in kidney tissue of STZ induced diabetic rats after 28 days of treatment. In-vitro advanced glycation end product inhibitory assay showed that AE possesses AGE inhibitory activity which can play important role in the prevention of DN progression. Present studies proved the potential of aqueous extract of Bauhinia variegata leaves and open a new alternative treatment approach for diabetic nephropathy.

Morin alleviates DSS-induced ulcerative colitis in mice via inhibition of inflammation and modulation of intestinal microbiota

Ulcerative colitis (UC) is a chronic inflammatory condition with recurrent and challenging symptoms. Effective treatments are lacking, making UC management a critical research area. Morin (MO), a flavonoid from the Moraceae family, shows potential as an anti-UC agent, but its mechanisms are not fully understood. Using a dextran sulfate sodium (DSS)-induced UC mouse model, we employed network pharmacology to predict MO’s therapeutic effects. Assessments included changes in body weight, disease activity index (DAI), and colon length. Immunofluorescence, hematoxylin and eosin (H&E), and PAS staining evaluated colon damage. ELISA and western blot analyzed inflammatory factors, tight junction (TJ)-associated proteins (Claudin-3, Occludin, ZO-1), and Mitogen-Activated Protein Kinase (MAPK)/ Nuclear Factor kappa B (NF-κB) pathways. 16S rRNA sequencing assessed gut microbiota diversity, confirmed by MO’s modulation via Fecal Microbial Transplantation (FMT). Early MO intervention reduced UC severity by improving weight, DAI scores, and colon length, increasing goblet cells, enhancing barrier function, and inhibiting MAPK/NF-κB pathways. MO enriched gut microbiota, favoring beneficial bacteria like Muribaculaceae and Erysipelotrichaceae while reducing harmful Erysipelotrichaceae and Muribaculaceae. This study highlights MO’s potential in UC management through inflammation control, mucosal integrity maintenance, and gut flora modulation.

Paneth cell differentiation associated with neoadjuvant therapy in esophageal adenocarcinoma.

Paneth cells and Paneth cell metaplasia are well-known in pathology as foundational components in the gastrointestinal system. When within malignant cells (Paneth cell differentiation [PCD]), however, the function and significance of these cells is less well understood. Here, we present findings from the first study focused on PCD in postneoadjuvant esophageal adenocarcinoma (EAC) resection specimens. Patients with EAC treated with neoadjuvant chemoradioation and followed by esophagectomy between 2012 and 2018 in our institution were retrospectively evaluated. A tissue microarray was constructed, and special and immunohistochemical stains were performed. A total of 64 cases were collected, of which 8 had PCD, as highlighted by periodic acid-Schiff with diastase staining. Adenocarcinomas with PCD were more commonly seen in patients 60 to 70 years of age and typically had a poorly differentiated morphology, observationally fewer stromal mucinous changes, and less lymph node metastasis. β-catenin activation induced by neoadjuvant therapy was more frequent in the PCD-positive cases. Patients with PCD-positive disease had low programmed cell death 1 ligand 1 levels, no positive or equivocal ERBB2 (HER2) expression, and low CD8-positive T-cell infiltration; they were also mismatch repair proficient. Patients with PCD-positive disease showed a survival pattern inferior to that of patients with PCD-negative disease. When induced by neoadjuvant therapy in EAC, PCD is associated with high β-catenin activation, less expression of targetable biomarkers, and a potentially worse clinical prognosis.

Protopine-Type Alkaloids Alleviate Lipopolysaccharide-Induced Intestinal Inflammation and Modulate the Gut Microbiota in Mice.

In this study, we assessed the therapeutic effects of Macleaya cordata (Willd). R. Br.-derived protopine-type alkaloids (MPTAs) in a mouse model of lipopolysaccharide (LPS)-induced intestinal inflammation. The experimental design involved the allocation of mice into distinct groups, including a control group, a model group treated with 6 mg/kg LPS, a berberine group treated with 50 mg/kg berberine hydrochloride and low-, medium- and high-dose MPTA groups treated with 6, 12 and 24 mg/kg MPTAs, respectively. Histological analysis of the ileum, jejunum and duodenum was performed using Hematoxylin and Eosin (H&E) staining. Moreover, the quantification of intestinal goblet cells (GCs) was performed based on PAS staining. The serum levels of IL-1β, IL-6, IL-8 and TNF-α were quantified using an enzyme-linked immunosorbent assay (ELISA), while the mRNA levels of TLR4, NF-κB p65, NLRP3, IL-6 and IL-1β were assessed using quantitative PCR (qPCR). The protein levels of TLR4, Md-2, MyD88, NF-κB p65 and NLRP3 were determined using Western blotting. Furthermore, the 16S rDNA sequences of bacterial taxa were amplified and analysed to determine alterations in the gut microbiota of the mice following MPTA treatment. Different doses of MPTAs were found to elicit distinct therapeutic effects, leading to enhanced intestinal morphology and an increased abundance of intestinal GCs. A significant decrease was noted in the levels of pro-inflammatory cytokines (IL-1β, IL-6, IL-8 and TNF-α). Additionally, the protein levels of TLR4, MyD88, NLRP3 and p-p65/p65 were markedly reduced by MPTA treatment. Furthermore, 16S rDNA sequencing analysis revealed that the administration of 24 mg/kg MPTAs facilitated the restoration of microbial composition.

N-acetylcysteine combined with insulin therapy can reduce myocardial injury induced by type 1 diabetes through the endoplasmic reticulum pathway

With the development of Type 1 diabetes mellitus (T1DM), various complications can be caused. Hyperglycemia affects the microenvironment of cardiomyocytes, changes endoplasmic reticulum homeostasis, triggers unfolding protein response and eventually promotes myocardial apoptosis. However, insulin therapy alone cannot effectively combat the complications caused by T1DM. Forty adult beagles were randomly divided into five groups: control group, diabetes mellitus group, insulin group, insulin combined with NAC group, and NAC group. 24-hour blood glucose, 120-day blood glucose, 120-day body weight, and serum FMN content were observed, furthermore, hematoxylin-eosin staining, Periodic acid Schiff reagent staining, and Sirius red staining of the myocardium were evaluated. The protein expressions of GRP78, ATF6, IRE1, PERK, JNK, CHOP, caspase 3, Bcl2, and Bax were detected. Results of the pathological section of myocardial tissue indicated that insulin combined with NAC therapy could improve myocardial pathological injury and glycogen deposition. Additionally, insulin combined with NAC therapy down-regulates the expression of GRP78, ATF6, IRE1, PERK, JNK, CHOP, caspase3, and Bax. These findings suggest that NAC has a phylactic effect on myocardial injury in beagles with T1DM, and the mechanism may be related to the improvement of endoplasmic reticulum stress-induced apoptosis.

Juglone suppresses vasculogenic mimicry in glioma through inhibition of HuR-mediated VEGF-A expression

Vasculogenic mimicry (VM) serves as a vascular-like channel that provides important substances for tumor growth and is a primary factor in glioblastoma (GBM) drug resistance. Human Antigen R (HuR)—an mRNA-binding protein—is highly expressed in GBM, closely related to tumor progression, and deemed a potential drug target. Although some small-molecule compounds have been identified to disrupt HuR binding to target mRNA, they remain in the preclinical research stage, suggesting the need for further validation and development of HuR inhibitors. In our study, we aim to screen for potential HuR inhibitors and investigate their efficacy and molecular mechanisms in GBM. We employed the fluorescence polarization method to identify HuR inhibitors from a natural compound library, confirming the efficacy of juglone in effectively inhibiting the binding of HuR to AREVegf-a. Further validation of the binding of juglone to HuR at the protein level was conducted through electrophoretic mobility shift analysis, surface plasmon resonance, and molecular docking. Furthermore, juglone demonstrated inhibitory effects on glioma growth and VM formation in vitro and in vivo. Moreover, it was observed that juglone reversed epithelial-mesenchymal transition by inhibiting the VEGF-A/VEGFR2/AKT/SNAIL signaling pathway. Finally, we established the capability of juglone to target HuR in U251 cells through HuR knockdown, mRNA stability, and cell thermal shift assays. Therefore, this study identifies juglone as a novel HuR inhibitor, potentially offering promise as a lead compound for anti-VM therapy in GBM by targeting HuR.Abbreviations: AKT, protein kinase B; ARE, adenine-and uridine-rich elements; CETSA, cellular thermal shift assay; DMEM, Dulbecco’s modified Eagle’s medium; ELISA, enzyme linked immune sorbent assay; EMSA, electrophoretic mobility shift assay; EMT, epithelial mesenchymal transition; FP, fluorescence polarization; GBM, glioblastoma; HTS, high-throughput screening; HuR, human antigen R; IF, Immunofluorescence; PAS, periodic acid-Schiff; PI3K, phosphoinositide-3 kinase; qRT-PCR, quantitative real-time PCR; RRMs, RNA recognition motifs; SPR, surface plasmon resonance. TMZ, temozolomide; VM, vasculogenic mimicry; VEGF-A, Vascular endothelial growth factor-A; VEGFR2, Vascular endothelial growth factor receptor-2.

Neuregulin 4 Attenuates Podocyte Injury and Proteinuria in Part by Activating AMPK/mTOR-Mediated Autophagy in Mice.

In this study, we investigate the effect of neuregulin 4 (NRG4) on podocyte damage in a mouse model of diabetic nephropathy (DN) and we elucidate the underlying molecular mechanisms. In vivo experiments were conducted using a C57BL/6 mouse model of DN to determine the effect of NRG4 on proteinuria and podocyte injury, and in vitro experiments were performed with conditionally immortalized mouse podocytes treated with high glucose and NRG4 to assess the protective effects of NRG4 on podocyte injury. Autophagy-related protein levels and related signaling pathways were evaluated both in vivo and in vitro. The involvement of the adenosine monophosphate-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) pathway was detected using chloroquine or AMPK inhibitors. The results showed that the AMPK/mTOR pathway was involved in the protective roles of NRG4 against high glucose-mediated podocyte injury. Also, NRG4 significantly decreased albuminuria in DN mice. PAS staining indicated that NRG4 mitigated glomerular volume and mesangium expansion in DN mice. Consistently, western blot and RT-PCR analyses confirmed that NRG4 decreased the expression of pro-fibrotic molecules in the glomeruli of DN mice. The immunofluorescence results showed that NRG4 retained expression of podocin and nephrin, whereas transmission electron microscopy revealed that NRG4 alleviated podocyte injury. In DN mice, NRG4 decreased podocyte apoptosis and increased expression of nephrin and podocin, while decreasing the expression of desmin and HIF1α. Overall, NRG4 improved albuminuria, glomerulosclerosis, glomerulomegaly, and hypoxia in DN mice. The in vitro experiments showed that NRG4 inhibited HG-induced podocyte injury and apoptosis. Furthermore, autophagy of the glomeruli decreased in DN mice, but reactivated following NRG4 intervention. NRG4 intervention was found to partially activate autophagy via the AMPK/mTOR signaling pathway. Consequently, when the AMPK/mTOR pathway was suppressed or autophagy was inhibited, the beneficial effects of NRG4 intervention on podocyte injury were diminished. These results indicate that NRG4 intervention attenuates podocyte injury and apoptosis by promoting autophagy in the kidneys of DN mice, in part, by activating the AMPK/mTOR signaling pathway.

Age changes in the gastric mucosa of male albino rat: histological, immunohistochemical, histomorphometric and biochemical study.

Age related changes in the stomach are associated with alterations in the structure and secretory function of the gastric glands. The present study aimed to investigate histological, histomorphometric and biochemical changes in the gastric mucosa of rats with age. Eighty adult male albino rats were randomly divided into four age groups, 20 rats in each (prepubertal, adolescent, adult, and senile). The gastric specimens were subjected to light microscopic examination using haematoxylin and eosin, PAS and Masson's trichrome stains. Immunohistochemical staining for caspase-3 and inducible nitric oxide synthase (iNOS) was carried out. Measurement of superoxide dismutase (SOD), glutathione peroxidase (GPx) and malondialdehyde (MDA) activity in gastric tissue homogenates was performed using ELISA. Quantitative analysis of vascular endothelial growth factor (VEGF) gene expression was done by real-time polymerase chain reaction (PCR). Light microscopic examination of gastric mucosa of senile rats revealed distortion of gastric glands and erosions. Surface mucous cells, mucous neck cells, parietal and chief cells exhibited cytoplasmic destruction, nuclear degeneration, apoptosis and oxidative damage. There was a significant decrease in the mean gastric mucosal thickness, increase in collagen content and decrease in mucous content with the advance of age. These morphological changes were associated with a significant decrease in SOD and GPx activity and increase in MDA activity, in addition to decreased VEGF gene expression. Gastric mucosa of aged rats showed histological and immunohistochemical alterations. These changes were associated with oxidative stress, decreased antioxidant capacity and decreased angiogenesis.

Anti-Inflammatory and Immunoregulatory Effects of Xiaoqinglong Decoction on a Murine Model of Combined Allergic Rhinitis and Asthma Syndrome

Background Xiaoqinglong Decoction (XQLD) is a traditional Chinese medicine formula used for the treatment of allergic rhinitis and asthma, including combined allergic rhinitis and asthma syndrome (CARAS), due to its anti-inflammatory and anti-seizure properties. However, the pharmacological activities and the underlying molecular mechanisms of XQLD remain to be elucidated . Hence, we investigated the effects of XQLD against inflammation in an ovalbumin (OVA)-induced CARAS model in BALB/c mice. Methods BALB/c mice were sensitized by intraperitoneal injection and aerosol inhalation of OVA. XQLD or dexamethasone was administered by oral gavage prior to OVA challenge for 7 consecutive days. Specific airway resistance (sRAW) was evaluated 24 h after the final challenge with OVA. Enzyme-linked immunosorbent assay was used to measure the levels of serum inflammatory factors. The nasal mucosa and lung tissue were examined for general morphology and goblet hyperplasia using hematoxylin and eosin or periodic acid Schiff staining. Flow cytometry was employed to analyze the frequencies of helper T lymphocytes in peripheral blood. Immunohistochemistry was performed to determine the expression of transcription factors in helper T lymphocytes and γδ TCR. Results XQLD significantly ameliorated the symptoms of CARAS model mice, suppressed serum levels of interferon-γ, interleukin (IL)-2, IL-4, IL-5, IL-17, while increasing levels of IL-10, transforming growth factor-β. Epithelium impairment, cilia loss, eosinophil infiltration, and goblet cell metaplasia were significantly reduced in the nasal mucosa and lung tissue sections. XQLD restored the balance of Th1/Th2 and Th17/Treg cell frequency in peripheral blood. Furthermore, XQLD down-modulated the expression of GATA-binding protein 3 (GATA3), retinoic acid receptor-related orphan receptor γt, and γδ T cell receptor (TCR) in the nasal mucosa, along with GATA3 and γδ TCR in the lung tissue. Conclusions XQLD alleviated OVA-induced CARAS in BALB/c mice through its anti-allergic effects and modulation of the immune system.

Effects of heat stress and long photoperiod on the prostate of rats

Objective: To examine light and heat effects on the morphological, histological, and micrometric structure of the prostate of rats. Methods: Thirty adult male rats were divided into three groups. The control group was kept under 20 °C-22 °C and an artificial 12 h/12 h day/night cycle; the temperature group was under normal light and at (42±1) °C heat for 4 to 5 h daily, and the light group was exposed to 8 h/16 h day/night cycle with 20 °C-22 °C. Rats were weighed five times (at the beginning of the study and every seven days). Five milliliters (mL) of their peripheral blood were taken. The tissue staining was performed using the hematoxylin-eosin (H &amp; E) stain and periodic acid-Schiff (PAS). In the following, tissue and cellular reactions to the PAS were examined. Results: Folds were located entirely on the surface of the anterior lobe and periphery of the other lobes. The secretory units in the anterior lobe were more than the lateral lobe. A strong reaction of the secretory cells to the PAS was observed. Testosterone serum levels of the light group also significantly increased compared to the control group (P&lt;0.05). The most histometric changes of the lobes were established in the lateral lobes. Heat stress resulted in a significant decrease in testosterone levels and transformed prostate tissue. The epithelium and parenchyma to scaffold ratio in the temperature group decreased. Conclusions: Maximum and minimum changes in the ventral lobe happened under the ascent of temperature and light, respectively. The ventral lobe in the study of prostatic hyperplasia should be more considered.

The role of uncoupling protein 2 in experimental periodontitis-associated renal injury in rats.

This study aims to explore changes in uncoupling protein 2 (UCP2) in experimental periodontitis-associated renal injury induced by ligation and investigate the effect of UCP2 on renal injury induced by periodontitis. Twelve Wistar male rats were randomly divided into two groups: control and periodontitis groups. A periodontal model was built by ligating the maxillary first molars area with 0.2 mm orthodontic ligature wire. After 8 weeks, the intraoral condition of the rats was observed and periodontal clinical indices such as gingival bleeding index (BI), periodontal probing depth (PD), and tooth mobility (TM) were detected. The maxillary bone was scanned by Micro CT to observe the alveolar bone resorption. The tissue mineral density (TMD), bone mineral density (BMD), bone volume fraction (BV/TV), trabecular thickness (Tb.Th), trabecular bone separation (Tb.Sp) were recorded, and the distance from the enamel bone boundary to the alveolar crest (CEJ-ABC) of the maxillary first molar was measured. The oxidative stress indexes such as malondialdehyde, glutathione (GSH), and superoxide dismutase (SOD) were detected using frozen rat kidney tissue. The gene expression of UCP2, nuclear factor erythroid 2-related factor 2 (Nrf2), and peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α) was observed by quantitative real-time polymerase chain reaction (qRT-PCR) test. The gingival tissue of the rats was used for immunohistochemical staining to observe the expression of the UCP2 protein. The fixed rat kidney tissue was used for hematoxylin-eosin (HE), periodic acid-schiff (PAS), MitoSOX Red, JC-1, and immunohistochemical staining to observe the renal histopathology, the level of reactive oxygen species (ROS), the level of mitochondrial membrane potential, and the expression of UCP2, Nrf2, and PGC-1α protein. Rat serum was collected to detect renal function indices, namely, blood urea nitrogen (BUN), creatinine (Cre), and albumin (Alb). Compared with the control group, the periodontitis group showed red, swollen, and soft gingival tissue, with gingival probing bleeding, periodontal PD increased, tooth loosening, alveolar bone resorption, decreased TMD, BMD, BV/TV, and Tb.Th indices, and increased Tb.Sp index, CEJ-ABC, and gingival UCP2 protein expression. Compared with the control group, the levels of MDA and ROS in the kidney tissue of periodontitis rats and the gene and protein expression of UCP2 increased, and the levels of MMP, GSH, and SOD and the gene and protein expression of Nrf2 and PGC-1α decreased. Renal functional indices, namely, BUN, Cre, and Alb, were not significantly different between the two groups. UCP2 may play a role in renal injury induced by periodontitis through oxidative stress.

New Glycotoxin Inhibitor from Sesuvium sesuvioides Mitigates Symptoms of Insulin Resistance and Diabetes by Suppressing AGE-RAGE Axis in Skeletal Muscle.

The current study intended to investigate the role of new natural compounds derived from the Sesuvium sesuvioides plant in mitigating symptoms of diabetes and insulin resistance in the diabetic mice model. Anti-advanced glycation activity, insulin, and adiponectin were quantified by enzyme-linked immunosorbent assay (ELISA). Glucose uptake was performed using enzymatic fluorescence assay, and glycogen synthesis was measured using PAS staining. Gene and protein expression was assessed using real time PCR (RT-PCR), and immunoblotting and fluorescent microscopy, respectively. The new flavonoid glycoside eupalitin 3-O-α-L-rhamnopyranosyl-(1→2)-β-D-glucopyranoside 1 isolated from S. sesuvioides exhibited anti-AGE activity by reducing human glycated albumin in liver cells. In a diabetic mouse model treated with compound 1, we observed improved glucose tolerance, increased adiponectin levels, and decreased insulin resistance. We also observed alleviated AGEs induced reduction in glucose uptake and restored glycogen synthesis in the compound 1-treated diabetic mice muscles. Exploring the molecular mechanism of action in skeletal muscle tissue of diabetic mice, we found that 1 reduced AGE-induced reactive oxygen species and the inflammatory gene in the muscle of diabetic mice. Additionally, 1 exhibited these effects by reducing the gene and protein expression of receptor for advanced glycation end products (RAGE) and inhibiting protein kinase C (PKC) delta activation. This further led us to demonstrate that compound 1 reduced serine phosphorylation of IRS-1, thereby restoring insulin sensitivity. We conclude that a new flavonoid glycoside from S. sesuvioides could be a therapeutic target for the treatment of symptoms of insulin resistance and diabetes.

Sirt6 overexpression relieves ferroptosis and delays the progression of diabetic nephropathy via Nrf2/GPX4 pathway

Objective Sirt6, reactive oxygen species and ferroptosis may participate in the pathogenesis of Diabetic Nephropathy (DN). Exploring the relationship between Sirt6, oxidative stress, and ferroptosis provides new scientific ideas to DN. Methods Human podocytes were stimulated with 30 mM glucose and 5.5 mM glucose. The mice of db/db group were randomly divided into two groups:12 weeks and 16 weeks. Collect mouse blood and urine specimens and renal cortices for investigations. HE, Masson, PAS and immunohistochemical staining were used to observe pathological changes. Western blot, RT-qPCR and immunofluorescence staining were used to evaluate expression of relevant molecules. CCK8 method was introduced to observe cell viability. The changes of podocyte mitochondrial membrane potential and mitochondrial morphology in each group were determined by JC-1 staining and Mito-Tracker. Results The expression level of Sirt6, Nrf2, SLC7A11, HO1, SOD2 and GPX4 were reduced, while ACSL4 was increased in DN. Blood glucose, BUN, Scr, TG, T-CHO and 24h urine protein were upregulated, while ALB was reduced in diabetic group. The treatment of Ferrostatin-1 significantly improved these changes, which proved ferroptosis was involved in the development of DN. Overexpression of Sirt6 might ameliorate the oxidation irritable reaction and ferroptosis. Sirt6 plasmid transfection increased mitochondrial membrane potential and protected morphology and structure of mitochondria. The application of Sirt6 siRNA could aggravated the damage manifestations. Conclusion High glucose stimulation could decrease the antioxidant capacity and increase formation of ROS and lipid peroxidation. Sirt6 might alleviate HG-induced mitochondrial dysfunction, podocyte injury and ferroptosis through regulating Nrf2/GPX4 pathway.

Creation of an in vitro model of GM1 gangliosidosis by CRISPR/Cas9 knocking-out the GLB1 gene in SH-SY5Y human neuronal cell line.

GM1 gangliosidosis is one type of hereditary error of metabolism that occurs due to the absence or reduction of β-galactosidase enzyme content in the lysosome of cells, including neurons. In vitro, the use of neural cell lines could facilitate the study of this disease. By creating a cell model of GM1 gangliosidosis on the SH-SY5Y human nerve cell line, it is possible to understand the main role of this enzyme in breaking down lipid substrate and other pathophysiologic phenomena this disease. To knock-out the human GLB1 gene, guides targeting exons 14 and 16 of the GLB1 gene were designed using the CRISPOR and CHOP-CHOP websites, and high-efficiency guides were selected for cloning in the PX458 vector. After confirming the cloning, the vectors were transformed into DH5α bacteria and then the target vector was extracted and transfected into human nerve cells (SH-SY5Y cell line) by electroporation. After 48 h, GFP+ cells were sorted using the FACS technique and homozygous (compound heterozygous) single cells were isolated using the serial dilution method and sequencing was done to confirm them. Finally, gap PCR tests, X-gal and Periodic acid-Schiff (PAS) staining, and qPCR were used to confirm the knock-out of the human GLB1 gene. Additionally, RNA sequencing data analysis from existing data of the Gene Expression Omnibus (GEO) was used to find the correlation of GLB1 with other genes, and then the top correlated genes were tested for further evaluation of knock-out effects. The nonviral introduction of two guides targeting exons 14 and 16 of the GLB1 gene into SH-SY5Y cells led to the deletion of a large fragment with a size of 4.62 kb. In contrast to the non-transfected cell, X-gal staining resulted in no blue color in GLB1 gene knock-out cells indicating the absence of β-galactosidase enzyme activity in these cells. Real-time PCR (qPCR) results confirmed the RNA-Seq analysis outcomes on the GEO data set and following the GLB1 gene knock-out, the expression of its downstream genes, NEU1 and CTSA, has been decreased. It has been also shown that the downregulation of GLB1-NEU1-CTSA complex gene was involved in suppressed proliferation and invasion ability of knock-out cells. This study proved that using dual guide RNA can be used as a simple and efficient tool for targeting the GLB1 gene in nerve cells and the knockout SH-SY5Y cells can be used as a model investigation of basic and therapeutic surveys for GM1 gangliosidosis disease.