Histopathological Observations Research Articles

Ameliorative effects of Turbinaria ornata extract on hepatocellular carcinoma induced by diethylnitrosamine in-vivo.

Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related deaths and the fifth most common cancer worldwide. Brown algae appeared to be a rich source of efficient and safe agents against many life-threatening diseases like cancer. Thus, the scope of this study was to investigate the therapeutic effects of Turbinaria ornata against experimentally induced HCC in a rat model. Accordingly, forty male albino rats were divided into four groups. HCC was induced by intraperitoneal injection with diethylnitrosamine (DENA) followed by carbon tetrachloride (CCL4). After four weeks of DENA + CCL4 injection and two weeks of treatment with Turbinaria ornata, rats were sacrificed to collect hepatic tissue and blood samples for histopathological observations and various biochemical markers such as aspartate aminotransferase (AST), alanine aminotransferase (ALT), alpha-fetoprotein (AFP), urea, creatinine, albumin (ALB), and alkaline phosphatase (ALP). Rats that were injected for four weeks with DENA + CCL4 showed a significant increase in AFP levels, transforming growth factor-beta (TGF-β) and tumor necrosis factor-alpha (TNF-α), as well as a high percentage of malignant changes in hepatic tissues. The extension of malignant changes in the rat liver tissues was markedly reduced using Turbinaria ornata, as the treatment displayed liver patterns similar to that of the normal control rats. Furthermore, rats with HCC fed with Turbinaria ornata extract for two weeks showed decreasing levels of TGF-β and TNF-α. These findings demonstrate that Turbinaria ornata supplement can prevent HCC development in hepatic rats; however, the exact mechanism requires further investigation.

Expression and clinical significance of PIKFYVE gene in hepatocellular carcinoma analyzed based on TCGA database and experimental validation

Objective: To investigate the expression and clinical significance of human FYVE finger-containing phosphoinositide kinase (PIKFYVE) in hepatocellular carcinoma (HCC) on the basis of cancer genome atlas (The cancer genome atlas, TCGA) database analysis and clinical samples experimental validation. Methods: Based on the data information of 424 clinical samples (including 374 cases of HCC tissues and 50 cases of nontumorous liver tissues) in the TCGA database, Cox regression analysis and Kaplan-Meier method were used to analyse the relationship between the PIKFYVE mRNA expression and the clinical characteristics, prognosis for survival of HCC patients. The relationship between the PIKFYVE gene and immune cell infiltration was examined by correlation analysis between the PIKFYVE gene and 24 immune cells. In addition, we analysed the correlation between the mRNA expression of PIKFYVE gene and RAC-alpha serine/threonine-protein kinase (AKT1), phosphatase and tensin homolog (PTEN), protein kinase C, alpha (PRKCA), inositol polyphosphate-5-phosphatase (INPP5D), phosphoinositide-3-kinase regulatory subunit 1(PIK3R1), Inositol Polyphosphate 4-phosphatase Type II (INPP4B) and phospholipase-C4 gene (PLCB4) in HCC tissues. Meanwhile, paraffin sections of highly differentiated, moderately differentiated, poorly differentiated, and nontumorous liver tissue in the Department of Pathology of the First Affiliated Hospital of Xinjiang Medical University were collected, each of which was 30 cases, and the histopathological observation was carried out by HE staining, and the expression levels of PIKFYVE and Ki67 proteins were verified by immunohistochemistry in each clinical sample. Results: The expression level of PIKFYVE gene in HCC tumours was significantly higher than that in normal liver tissues (P=0.000 2, P<0.01), and the overall survival of patients in the low PIKFYVE expression group was significantly longer than that in the high expression group (HR=1.57, 95%CI: 1.10～2.25, P=0.014). The results of Univariate Cox regression analysis showed that there was an effect of TNM stage, pathological stage, tumour status and residual tumour on Overall survival (OS) (P<0.05), and the expression level of PIKFYVE had an effect on OS survival (P<0.05); the PIKFYVE prognostic risk model score ratio was HR=1.533 (1.077-2.181, P=0.018). Multivariate Cox regression analysis showed a PIKFYVE prognostic risk model score ratio HR=1.481 (0.886-2.476, P=0.134) and an area under the Receiver Operating Characteristic curve of 0.640, which was greater than 0.5, suggesting that the PIKFYVE prognostic risk model has a predictive value in survival prediction. Correlation analysis showed that the expression level of PIKFYVE was highly correlated with immune cell infiltration and TP53 (P<0.01). The immunohistochemistry staining results showed that the expression of PIKFYVE in HCC tissues was significantly higher than that of nontumorous tissues (P<0.05), and there was a negative correlation with the degree of differentiation. Conclusion: PIKFYVE, as an independent risk factor for HCC, is expected to be developed as a clinical diagnostic biomarker for HCC, which will provide a reference for new drugs for the treatment of HCC.

Design of Nanocrystalline Suspension of Dutasteride for Intramuscular Prolonged Delivery.

The aim of the study is to formulate an injectable nanocrystalline suspension (NS) of dutasteride (DTS), a hydrophobic 5α-reductase inhibitor used to treat benign prostatic hyperplasia and scalp hair loss, for parenteral long-acting delivery. A DTS-loaded NS (DTS-NS, 40 mg/mL DTS) was prepared using a lab-scale bead-milling technique. The optimized DTS-NS prepared using Tween 80 (0.5% w/v) as a nano-suspending agent, was characterized as follows: rod/rectangular shape; particle size of 324 nm; zeta potential of -11 mV; and decreased drug crystallinity compared with intact drug powder. The DTS-NS exhibited a markedly protracted drug concentration-time profile following intramuscular injection, reaching a maximum concentration after 8.40 days, with an elimination half-life of 9.94 days in rats. Histopathological observations revealed a granulomatous inflammatory response at the injection site 7 days after intramuscular administration, which significantly subsided by day 14 and showed minimal inflammation by day 28. These findings suggest that the nanosuspension system is a promising approach for the sustained release parenteral DTS delivery, with a protracted pharmacokinetic profile and tolerable local inflammation.

An ultrasound-activated piezoelectric sonosensitizer enhances mitochondrial depolarization for effective treatment of orthotopic glioma

Despite the significant advancements in piezoelectric materials for sonodynamic therapy (SDT), the suppression of orthotopic glioma remains challenging, primarily due to the unclear mechanism and the restriction of blood-brain barrier (BBB). Herein, we proposed that layered piezoelectric SrBi2Ta2O9 nanoparticles (SBTO NPs) could effectively depolarize the mitochondrial membrane potential (ΔΨm) of glioma cells under ultrasound (US) exposure. The US-induced band bending in SBTO NPs enhanced redox ability, promoting an increase in reactive oxygen species (ROS) generation. The in vitro results proved that SBTO NPs selectively accumulated in mitochondria under US and induced apoptosis in a mitochondrial depolarization manner mediated by the generation of ROS and free charges. Furthermore, SBTO NPs could cross the BBB and then accumulate in gliomas through US/microbubbles (MBs) procedure and protein-mediated transport. The therapeutic effect of piezoelectric SBTO NPs mediated SDT was proved in the orthotopic glioma mouse model. As validated by the histopathological observation and the long-term evaluation, the good biocompatibility and biosafety of SBTO NPs make it possible for deep tumor therapy, and worthy for further preclinical study. Statement of significanceEmploying piezoelectric sonosensitizers for sonodynamic therapy (SDT) has emerged as a promising strategy for cancer treatment; however, the unclear mechanism and blood-brain barrier (BBB) limit the effectiveness of SDT in glioma. Herein, we developed piezoelectric SrBi2Ta2O9 nanoparticles (SBTO NPs) with a built-in electric field for glioma treatment and explored the underlying therapeutic mechanism. Notably, SBTO NPs selectively accumulated in mitochondria under ultrasound (US) and induced apoptosis in a mitochondrial depolarization manner, which is mediated by the generation of reactive oxygen species (ROS) and free charges. In an orthotopic glioma mouse model, SBTO NPs were delivered into the glioma through US/microbubbles and transferrin-mediated transport pathways, inhibiting tumor growth. This work provides a new paradigm for the treatment of orthotopic glioma and other tumor types.

Construction and characterization of different hemolysin gene deletion strains in Vibrio parahaemolyticus (ΔhlyA, ΔhlyIII) and evaluation of their virulence

Vibrio parahaemolyticus, a halophilic food-borne pathogen, possesses an arsenal of virulence factors. The pathogenicity of V. parahaemolyticus results from a combination of various virulence factors. HlyA and hlyIII genes are presumed to function in hemolysis, in addition to tdh and trh in V. parahaemolyticus. To confirm the hemolytic function of genes hlyA and hlyIII, ΔhlyA and ΔhlyIII strains of V. parahaemolyticus were separately constructed via homologous recombination. The cytotoxicity and pathogenicity of the ΔhlyA and ΔhlyIII strains were evaluated using a Tetrahymena-Vibrio co-culture model and an immersion challenge in Litopenaeus vannamei. Results indicated that the hemolytic activity of the ΔhlyA and ΔhlyIII strains decreased by approximately 31.4 % and 24.9 % respectively, compared to the WT strain. Both ΔhlyA and ΔhlyIII exhibited reduced cytotoxicity towards Tetrahymena. Then shrimp infection experiments showed LD50 values for ΔhlyA and ΔhlyIII of 3.06 × 108 CFU/mL and 1.23 × 108 CFU/mL, respectively, both higher than the WT strain’s value of 2.57 × 107 CFU/mL. Histopathological observations revealed that hepatopancreas from shrimps challenged with ΔhlyA and ΔhlyIII exhibited mild symptoms, whereas those challenged with the WT strain displayed severe AHPND. These findings indicate that the ΔhlyA and ΔhlyIII strains are significantly less virulent than the WT strain. In conclusion, both hlyA and hlyIII are vital virulence genes involved in hemolytic and cytotoxic of V. parahaemolyticus.

Growth performances of Clarias gariepinus juveniles fed with Jatropha curcas seed meal

Jatropha curcas is an oil producing seed with high nutritional qualities for consumption. However, due to inherent anti-nutritional factors, appropriate processing is necessary to improve its nutrients utilization. This study was aimed at processing J. curcas using solvent-extraction method and evaluating its inclusion effects in the diet for Clarias gariepinus on its growth status. J. curcas meal was extracted using 80 % methanol to remove phorbol esters and other anti-nutrients. Thereafter, five iso-nitrogenous diets (40 % crude protein) were formulated containing J. curcas meal replacements of soybeans meal at 0 % (TRT0), 25 % (TRT25), 50 % (TRT50), 75 % (TRT75) and 100 % (TRT100) and fed to C. gariepinus for 70 days. Results showed that TRT25 fish had the highest mean weight gain and specific growth. The serum alkaline phosphate, alanine aminotransferase, and aspartate aminotransferase levels began to rise significantly (p ≤ 0.05) at 50 % replacement. Likewise, the assessment of gut ecology and morphology indicated that TRT25 had significantly (p ≤ 0.05) the highest gut bacteria colony forming unit (2.60 ×104±0.02cfu/g) and most favorable area of absorption (0.16±0.01 cm2). The histopathological observation of the fish intestine, liver and gills indicated no visible deformity in TRT0 and TRT25 fish. However, various degrees of degenerations were observed in the fish fed with 50 % and higher inclusion of J. curcas. This study showed that methanol-extracted J. curcas meal can be effectively utilized for C. gariepinus at 25 % of replacement.

The triple combination DBDx alleviates cytokine storm and related lung injury

Cytokine storm is a life-threatening disorder, and therapeutic treatments are urgently needed. Here, we investigated the anti-cytokine storm efficacy of DBDx, a triple drug combination composed of dipyridamole, ubenimex and dexamethasone. Evaluated by lipopolysaccharide (LPS)-induced cytokine storm murine model, DBDx significantly improved survival rate and prolonged survival time of the model mice. Notably, the efficacy of DBDx was higher than that of dipyridamole, ubenimex and dexamethasone. Determined by ELISA, DBDx significantly reduced the LPS-stimulated serum levels of TNF-α, IL-6 and IL-1β in mice. Luminex assay showed that DBDx suppressed the serum levels of a wide variety of inflammatory cytokines and chemokines, which was more potent than dexamethasone alone. Otherwise, DBDx exerted similar inhibitory effects on cytokine profiles in bronchoalveolar lavage fluid. Histopathological observation showed that DBDx significantly reduced the LPS-induced thickening of alveolar septum, indicating its suppression of capillary congestion, edema and neutrophil infiltration in the lung. Ultra-structure analysis showed that DBDx suppressed the LPS-induced morphological changes of microvilli in type II pneumocytes. In vitro experiment showed that DBDx inhibited IL-6 and TNF-α secretion in THP-1 cells, and downregulated TLR4/NF-κB/HIF-1α signaling pathway. All of these results demonstrate that DBDx, a triple combination of clinical orally-administered drugs, can alleviate cytokine storm and related lung injury. DBDx is beneficial for treating cytokine storm disorders.

Molecular characterization and immunopathological investigation of Avian reticuloendotheliosis virus in breeder flocks in Egypt

BackgroundReticuloendotheliosis virus (REV) is an oncogenic immunosuppressive retrovirus that infects different kinds of avian species; posing significant economic losses to the poultry industry worldwide.MethodsIn Egypt, there is an unidentified disease associated with the runting-stunting syndrome with neoplasia, suspected to be REV, that has been continuously monitored in several breeder flocks. To diagnose and analyze REV by cell cultures, enzyme-linked immunosorbent assay (ELISA), histopathological investigation, the polymerase chain reaction (PCR) test, and sequencing analysis, 200 blood samples, and 50 tissue specimens were collected. The current study targets the occurrence and genetic characteristics of a viral neoplastic disease, resembling REV infection, circulating in breeder flocks from 2022 to 2023 in the Ismailia, El-Sharqia, and El-Dakahliya governorates.ResultHere, REV was isolated on chicken embryo fibroblast cell culture; exhibiting cell aggregation, rounding, and cell detachments. Collectively, only 70 serum samples were positive for anti‐REV antibodies with seroprevalence rates of 35% based on the ELISA test. The histopathological observation demonstrated lymphoreticular tumors in the liver, spleen, and other examined organs. The immunohistochemical staining method confirmed the REV-positive signals in all examined organs (liver, kidney, spleen, bursa, ovaries) except for the heart. The PCR assay of the LTR gene assessed 370 base pairs with only 5 positive samples with a percentage of 16.6%. Three positive samples were further sequenced and submitted to the Genbank under accession numbers (PP763709, PP763710, PP763711). Phylogenetic analysis of the REV-LTR gene showed that our three isolates (Sharquia-1-REV, Ismilia-2-REV, Mansoura-3-REV) are REV subtype III which predominantly circulated in breeders in Egypt. These three isolates are highest similar to American, Chinese, and Taiwanese REV reference strains, and other Egyptian strains with nucleotide identity percentages of 100%, 99%, and 99%; respectively, and on the amino acid identity level were with (99–100%), (98%, 99%), (99%, 100%); respectively.ConclusionsThis study established that REV infection was extensively distributed in the breeders and became one of the causes of the clinical outbreaks of tumors, raising awareness of REV as the causative agent of avian oncogenic disease in Egypt.

Elucidation of the Possible Molecular Mechanism for the Anti-Apoptotic Effect of Terpene-Rich Extract of Amomum subulatum and Coriandrum sativum against Rodent Model of Cardiotoxicity

Background: Spices, including Amomum subulatum and Coriander sativum, are not only popular for improving food flavour but are also accepted worldwide for their health benefits in various disorders. Objective: This study aimed to elucidate the possible mechanism for the anti-apoptotic effect of terpene-rich extract of Amomum subulatum and Coriandrum sativum against the rodent model of cardiotoxicity. Methods: Effects of standardized terpenoids-rich aqueous methanolic extract of two commonly used spices, viz. Amomum subulatum Roxb. and Coriandrum sativum Linn., were investigated on doxorubicin-induced apoptotic changes and cardiotoxicity in Wistar rats with the aim of in-vestigating the mechanism. Prior to the in vivo experiment, the extracts were subjected to quan-titative estimation of possible bioactive markers of the terpenes by employing a newly devel-oped, optimized, and validated GC-MS method along with TLC profiling. Results: Cardiotoxicity was evident from elevated creatinine kinase (CK-MB), lactate dehydro-genase (LDH), and malondialdehyde (MDA) levels in the toxic control group after treatment with doxorubicin (2.5mg/kg i.p. given twice a week for three weeks). Caspase-3, Tumor necrosis factor-α (TNF-α), and Interleukin-6 (IL-6) were also induced in animals treated with doxorubi-cin. Treatment with Amomum subulatum and Coriandrum sativum at the doses of 100 and 200mg/kg exhibited significant (P&lt;0.001) reversal of CK-MB, LDH, MDA, Caspase-3, TNF-α, and IL-6 levels. This protective effect was further supported by the results of DNA gel electro-phoresis and histopathological observations. Conclusion: This study supports the cardioprotective role of selected spices against doxorubicin-induced cardiotoxicity through the anti-apoptotic mechanism.

Effect of co-exposure to multiple metals (Pb, Cd, Cr, Hg, Fe, Mn and Ni) and metalloid (As) on liver function in Swiss albino mice.

The study examined the cumulative toxic effect of multiple elements, As, Pb, Cd, Cr, Hg, Fe, Mn and Ni on the liver function and their association with inflammation and apoptosis. To explore the health consequence of simultaneous exposure to multiple metals and metalloid, male and female Swiss Albino mice were randomly divided into 14 groups and subjected to different doses [MPL (maximum permissible limit), 1×, 5×, 10×, 50× or 100×] of metal(loid)s mixture via drinking water for 8weeks. Data showed that combined effect of multiple elements impaired the liver function. This was associated with significant decrease in the antioxidant enzymes and the elevation in lipid peroxidation for high exposure dose of 50× and 100× (p < 0.05). The metal(loid)s mixture exposure led to significant increase (p < 0.05) in cytokines, TNF-α, IL-6 and effector caspases (3 and 6) in exposure groups above 10× dose. Histopathological observation also revealed significant damage in the hepatic tissue on exposure to high dose. Dose dependent accumulation of respective elements (As, Cd, Cr, and Pb) in the liver was observed in each of the exposure groups. However, similar dose related increment was not observed for essential metals such as Ni, Fe and Mn. Differential accumulation of metals in the liver may be attributed to the effect of co-contaminant exposure, which could affect the divalent cation absorption due to antagonism and competitive transport process. Overall findings in this study manifest the complexity of possible joint effect of co-exposure to multiple metals and metalloid on the liver function.

Lactococcus garvieae as a Novel Pathogen in Cultured Pufferfish (Takifugu obscurus) in China

In October 2023, a disease outbreak in pufferfish (Takifugu obscurus) farms in Zhongshan City, Guangdong, China, caused high mortality. Diseased fish (mean length: 15 ± 1 cm) exhibited swimming disorders, fin rot, hemorrhage, and an enlarged spleen. Histopathological observations generally revealed inflammation, necrosis, and congestion in the spleen, kidneys, and brain tissues. The most severe pathological changes included interstitial edema and tubular atrophy in the kidneys, hemosiderin deposition in the spleen, massive red blood cell infiltration, and a decrease in lymphocytes. A single strain of bacteria (Tol-1) was isolated from the diseased pufferfish and identified as a Gram-positive streptococcus strain, exhibiting α-hemolysis on sheep blood agar plates. Through biochemical characterization, 16S rDNA sequencing, morphological analysis, and specific primer-based identification, the Tol-1 strain was identified as Lactococcus garvieae, serotype I. Antimicrobial susceptibility testing indicated that Tol-1 was sensitive to Chloramphenicol, Ampicillin, Cephalexin, and Doxycycline, but resistant to Kanamycin, Gentamicin and Ciprofloxacin. In addition, 15 common virulence factors were detected in the Tol-1 strain, including adhPav, adhPsaA, adhC I–II, adh, and hly 1–3. Pufferfish (mean length: 17 ± 1 cm) subjected to artificial infection via intraperitoneal injection (IP) with the Tol-1 strain exhibited clinical symptoms and histopathological damage similar to those observed in naturally infected fish. An infection dose of 1 × 105 CFU/fish resulted in 80% mortality. The study fulfilled Koch’s postulates, indicating that the disease outbreak in pufferfish was caused by L. garvieae, which exhibited a high mortality rate in pufferfish despite the subtle clinical symptoms. These results serve as a warning for pufferfish farming areas and provide a scientific basis for future prevention and control efforts.

Untargeted Metabolite Profiling Reveals Acute Toxicity of Pentosidine on Adipose Tissue of Rats.

Background: Pentosidine is an advanced glycation end product that is commonly found in heat-processed foods. Pentosidine has been involved in the occurrence and development of some chronic diseases. It was reported that pentosidine exposure can impair the function of the liver and kidneys. Adipose tissue, as an active endocrine organ, plays an important role in maintaining the normal physiological function of cells. However, the metabolic mechanism that causes pentosidine to induce toxicity in adipose tissue remains unclear. Methods: In the study, thirty male Sprague-Dawley rats were divided into a normal diet group, low dose group, and high dose group. A non-targeted metabolomics approach was used to compare the metabolic profiles of adipose tissue between the pentosidine and normal diet groups. Furthermore, histopathological observation and body weight change analysis were performed to test the results of the metabolomics analysis. Results: A total of forty-two differential metabolites were identified. Pentosidine mainly disturbed twelve metabolic pathways, such as ascorbate and aldarate metabolism, glycine, serine, and threonine metabolism, sulfur metabolism, pyruvate metabolism, etc. Additionally, pyruvic acid was identified as a possible key upregulated metabolite involved in thirty-four metabolic pathways. α-Ketoglutaric acid was named as a probable key downregulated metabolite involved in nineteen metabolic pathways based on enrichment network analysis. In addition, histopathological analysis and body weight changes confirmed the results of the metabolomics analysis. Conclusions: These results provided a new perspective for the molecular mechanisms of adipose tissue toxicity induced by pentosidine.

Chemoprotective Effect of Eugenol on Repeated dose Doxorubicin, Induced Genotoxic Lesions in Mouse Bone Marrow

In present study, an attempt has been made to evaluate the possible in vivo chemoprotective potential of eugenol against doxorubicin induced genotoxic lesion in bone marrow. Swiss albino female mice were administered Eugenol (10 mg/kg/day, orally) 15 days before doxorubicin administration as well as concomitantly with doxorubicin. Genotoxic lesions were induced by a repeated dose intraperitoneal injection of doxorubicin (5mg/kg b.w. i.p. upto 9 days (alternate day). Micronuclei assay and COMET assay were carried out to determine genetic damage in bone marrow. Histopathological evaluation of bone marrow was also done to study lesions at cellular level. The study results demonstrated that eugenol significantly mitigated Doxorubicin induced micronuclei formation and DNA damage in the bone marrow niche. Histopathological observations revealed that Doxorubicn-intoxication resulted in massive structural impairment of bone marrow which was reduced by eugenol administration. Amelioration by eugenol is more significant in mice those received eugenol 15 days prior to Doxorubicin administration suggesting its chemoprotection. The present study suggests eugenol has the promising chemoprotective effect against Doxorubin-induced genetic lesions in mice bone marrow.

Role of toll-like receptor 4/mutant myeloid differentiation primary response 88/nuclear factor kappa-B mediated inflammation in diabetes mellitus with Northwest dryness syndrome.

To investigate the role of toll-like receptor 4 (TLR4)/mutant myeloid differentiation primary response 88 (MyD88)/nuclear factor kappa-B (NF-κB) signaling pathway-mediated inflammation in diabetes mellitus with Northwest dryness syndrome. Rats were randomly divided into the normal control, type 2 diabetes (T2DM) model, Northwest dryness syndrome + T2DM (Northwest dryness), and simple internal dampness + T2DM (internal dampness) groups. Enzyme-linked immunosorbent assay was used to detect biochemical indexes and inflammatory factors. The histopathological observation was performed. Quantitative real-time polymerase chain reaction and Western blot analysis were used to detect the mRNA and protein expression levels, respectively. Compared with the T2DM group, the glycosylated hemoglobin A1c, insulin, glucose tolerance, the homeostasis model assessment of insulin resistance, tumor necrosis factor-α, interleukin 1β, interleukin 16, malondialdehyde, blood lipid, alanine aminotransferase, and aspartate aminotransferase were significantly elevated in the internal dampness group. Their levels were significantly elevated in the Northwest dryness group than in the T2DM and internal dampness groups. The superoxide dismutase, glutathione peroxidase, liver glycogen, and organ-to-weight ratio were significantly declined in the internal dampness group and the Northwest dryness group than in the T2DM group. However, these levels were elevated in the Northwest dryness group than in the internal dampness group. Moreover, the mRNA expression levels of interferon regulatory factor 5 and NF-κB p65, and the protein expression levels of TLR4, MyD88, and NF-κB were significantly higher in the internal dampness and the Northwest dryness groups than the T2DM group. Additionally, the mRNA and protein levels were significantly higher in the Northwest dryness group than in the internal dampness group. Northwest dryness syndrome-mediated TLR4/MyD88/NF-κB pathway and chronic inflammation might be associated with the occurrence and development of T2DM.

Qingfei Huoxue Decoction and Its Active Component Narirutin Alleviate LPS-Induced Acute Lung Injury by Regulating TLR4/NF-κB Pathway Mediated Inflammation.

Acute lung injury (ALI) is a life-threatening clinical syndrome with high mortality. Currently, the safe and effective therapies for ALI patients are still limited. Qingfei Huoxue decoction (QFHXD) is a hospital agreement prescription for treating pulmonary diseases and displays a remarkable efficacy. However, the pharmacological effect of QFHXD on preventing lipopolysaccharide (LPS)-induced ALI has yet to be reported, let alone questions of potential molecular mechanisms and anti-ALI active substances. To answer the above-mentioned questions, histopathological observation and kit detection were performed to estimate the protective effect of QFHXD pretreatment against LPS-induced ALI. Based on comprehensive chemical profiling of QFHXD, a network pharmacology strategy and experimental validation were integrated to elucidate the underlying functional mechanisms. The potential anti-ALI active components were identified by molecular docking. The anti-ALI activity of narirutin and its anti-inflammatory mechanism were further validated using animal and molecular experiments. Pretreatment with different doses of QFHXD effectively mitigated histopathological lesions and systemic inflammation caused by LPS stimulation. A detailed analysis of established compound-target-disease network revealed the strong correlation between anti-ALI action of QFHXD and inflammatory mechanisms. Compared with the model group, QFHXD intervention markedly restrained the abnormally increased transcription and protein levels of pro-inflammatory factors (TLR4, NF-κB, IL-6, IL-1β, and TNF-α) in lung tissues of ALI mice. The results of molecular docking highlighted the anti-ALI potential of narirutin targeting to TLR4 and NF-κB p65. In addition to the protective effect of narirutin on suppressing LPS-induced pathological changes, we found that narirutin pretreatment effectively normalized the disordered protein levels of above pro-inflammatory factors of ALI mice. These interesting findings indicate the beneficial effects of QFHXD and its active component narirutin against ALI partly via regulating TLR4/NF-κB mediated inflammation. This work contributes to the development of novel medications for ALI patients.

Integrated gut microbiota and serum pharmacochemistry reveal the mechanisms of wine steaming in alleviating rhubarb diarrhea

BackgroundLong-term use of rhubarb (RH) can cause adverse gastrointestinal reactions (such as diarrhea), whereas RH steaming with wine (PRH) can alleviate RH-induced diarrhea. However, the potential material basis and mechanisms by which wine steaming alleviates diarrhea caused by RH remain unclear. PurposeTo reveal the potential material basis and underlying mechanisms of wine steaming in alleviating diarrhea caused by RH from the perspective of small intestinal flora and immune function. MethodsThe major anthraquinone/anthrone components were detected using high-performance liquid chromatography (HPLC). Constipation model mice were replicated using loperamide hydrochloride and were administered RH and PRH for six consecutive weeks. Histopathological observation (duodenum, jejunum, and ileum) was performed using hematoxylin-eosin (HE) staining, and the serum levels of inflammatory cytokines, immunoglobulin G (IgG), and immunoglobulin A (IgA) were examined. CD4+, CD8+, and Treg cells counts in peripheral blood were determined using flow cytometry; The protein expression of Toll-like receptor 4 (TLR4) and nuclear factor kappa-B (NF-κB) was determined using immunohistochemistry (IHC) and western blot (WB). The small intestine contents and feces were analyzed by 16 S rRNA sequencing and the contents of short chain fatty acids (SCFAs) in feces were determined using gas chromatography-mass spectrometry (GC–MS). Ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) was used to analyze the blood absorption compounds and endogenous metabolites. ResultsThe levels of the major anthraquinone/anthrone components were decreased in PRH. RH and PRH both increased the wet fecal weight at 12 h (WFW-12) and fecal water rate (FWR), alleviated the dry and black fecal morphology, and relieved small intestine injuries in the second week. In the fourth week, although RH and PRH alleviated the abnormal levels of indicators in the model mice (fecal water rate, immune cells percentage, and TLR4/NF-κB expression), minor small intestinal damage was observed. Compared to that at the fourth week, RH and PRH increased the levels of WFW-12, FWR, inflammatory cytokines, and TLR4/NF-κB expression, and decreased the levels of IgG/IgA and immune cells with extended administration (sixth week). Further, damage to the small intestine worsened (severe ileal damage) and different degrees of loose stools were observed in RH- and PRH-administered mice in the sixth week. Compared with those in the control group, the levels of WFW-12, FWR, inflammatory cytokines, TLR4/NF-κB expression, IgG/IgA, and immune cell percentage were significantly different in the RH-H and PRH-H mice at the sixth week (except for CD8+in PRH-H). Further, RH and PRH disturbed the gut microbiota (GM) (Lactobacillus and Dubosiella decreased, Aerococcus and Corynebacterium increased) and obviously reduced the content of SCFAs (acetic acid, butyric acid, and isobutyric acid). However, almost all the results indicated a lower impact of PRH than that of RH. Metabolic pathways mainly involved in glycerophospholipid metabolism were identified along with a total of 21 blood absorption components, including anthraquinones, anthrones, flavanols, and tannins. The correlation analysis showed a positive correlation of pathogenic bacteria (Aerococcus and Corynebacterium) with inflammatory cytokines, TLR4/NF-κB, LysoPC(20:0/0:0), and PE (16:0/20:4(8Z,11Z,14Z,17Z)) and a negative correlation with immune cells and SCFAs (acetic acid and isobutyric acid); however, the opposite results were observed for beneficial bacteria (Lactobacillus and Dubosiella). ConclusionOverall, PRH can alleviate RH-induced diarrhea by recovering the GM imbalance and abnormal levels of GM-mediated SCFAs, alleviating the decrease in cellular immune function and abnormal expression of TLR4/NF-κB, thereby suppressing the release of inflammatory factors, possibly, through its lower content of anthraquinones. This study explored for the first time the processing mechanism of wine steaming in alleviating RH-induced diarrhea from the aspects of small intestinal flora and small intestinal immune function.

Zinc Sulfate and α-tocopherol Supplementation Enhance Reproductive Performance in Male Albino Rats (Rattus norvegicus) With Lead Acetate Toxicity

Metal toxicity from lead affects reproductive organ function by activating reactive oxygen species processes. This study aims to see how α-tocopherol and zinc sulfate (ZnSO4) affect gonads, liver, follicle-stimulating hormone, luteinizing hormone, spermatogenesis (the amount of spermatogonia, spermatocytes, and spermatids), and Leydig cells in male albino rats (Rattus norvegicus) exposed to lead acetate Pb(CH3COO)2. The samples used were 25 male Wistar rats aged 4 months, separated into five groups. For 30 days, all treatment groups were exposed to Pb(CH3COO)2 at a level of 50-mg/kg body weight (BW). The T1 group was given a dosage of 100-mg/kg BW of α-tocopherol. The ZnSO4 was given to the T2 group at a dose of 0.54-mg/kg BW. Meanwhile, the T3 group was given a mixture of ZnSO4 at 0.54-mg/kg BW and α-tocopherol at 100-mg/kg BW orally. ELISA test was carried out to determine the level of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) in a blood plasma sample of 100 μl / 1 mg. Histopathological observations made on the liver included counting damaged cells and seminiferous tubules that included counting the amount of spermatogonia, spermatocytes, spermatids, Sertoli cells, and Leydig cells. Using SPSS 20 software, the collected data were analyzed using analysis of variance, followed by Duncan’s test with a 95% simultaneous confidence level. The highest average levels of FSH and LH in the T3 group were 3.6162 mIU/mL and 14.9658 mIU/mL. The finding showed that Pb(CH3COO)2 caused disruptions in the spermatogenesis and Leydig cell processes. Exogenous antioxidants in combination with ZnSO4 and α-tocopherol had significant effect on enhancing reproductive performance in animals exposed to Pb(CH3COO)2.

Recoverability of zebrafish from decabromodiphenyl ether exposure: The persisted interference with extracellular matrix production and collagen synthesis and the enhancement of arrhythmias

As a widely used brominated flame retardant, the widespread presence of decabromodiphenyl ether (BDE-209) in the natural environment and the toxicity risks it poses are well established, but the recoverability of BDE-209-induced individual injuries remains unknown. Therefore, a 7-day depuration experiment following a 4-day exposure of zebrafish to BDE-209 was conducted to confirm the recoverability and its mode of action. Oxidative stress after depuration was significantly reduced compared with BDE-209 exposure as indicated by the decreased expression level of oxidative stress-related genes and the reduced MDA, Gpx, and GST in zebrafish, indicating a gradual recovery of antioxidant activity. However, BDE-209 inhibition of extracellular matrix (ECM) proteins worsened after depuration. Mechanistically, BDE-209 mediated ECM production and secretion by down-regulating integrin expression. Furthermore, BDE-209 inhibition of collagen synthesis worsened after depuration. Biochemical assays and histopathological observations revealed a same result in zebrafish. Mechanistically, lysine hydroxylation is inhibited thereby affecting collagen synthesis. Interestingly, zebrafish showed arrhythmia after depuration compared to BDE-209 exposure, and abnormal changes in ATPase levels indicated that disturbances in Ca2+ homeostasis contributed to arrhythmia. Collectively, BDE-209-induced interference with ECM production and collagen synthesis persisted after depuration, which will provide new insights for understanding the recovery patterns of individuals under BDE-209 stress.

Purified L-glutaminase effects against multidrug-resistant Pseudomonas aeruginosa in experimental vaginosis model: An immunological and histopathological observation

The antimicrobial activity of crude and purified L-glutaminase (EC 3.5.1.2), obtained from Lactobacillus gasseri, was evaluated against multidrug-resistant Pseudomonas aeruginosa in the in vivo vaginosis condition. The L-glutaminase possessed significant antimicrobial and anti-biofilm formation activity against multi-drug resistance P. aeruginosa, which were confirmed in the BALBc rat vaginosis model, together with its effects on the immunological and histopathological aspects. The untreated animals showed heavy vaginitis, characterized by sub-epithelial edema and infiltration of mononuclear leukocytes, perivascular heavy inflammatory cells infiltration in the vaginal tissue, and moderate stromal edema. However, the L-glutaminase treatment exhibited no changes in vaginal tissue structure with normal appearance of the epithelium and lamina propria with marked repair of the vaginal section when compared with normal, uninfected, control group A. The immunomodulatory actions of the L-glutaminase were confirmed by observance of higher concentrations of tumor necrosis factor-γ (TNF-γ), and interleukin −12 (IL-12) in treated animals, while the interleukin-10 (IL-10) was higher in the infected, untreated animals' sera samples. Therefore, the L-glutaminase showed corrective and healing actions, which were observed through histopathological observations of the vaginal tissue. The investigations led to imply that L-glutaminase may have the potential to be an effective antimicrobial agent for preventing and inhibiting bacterial growth, as well as inhibiting the biofilm formation in the P. aeruginosa-originated vaginosis. The observations may be of promising value for future clinical use.

Exploring the therapeutic potential of rabdoternin E in lung cancer treatment: Targeting the ROS/p38 MAPK/JNK signaling pathway.

Lung cancer has the highest incidence and mortality rates of all cancer types in China and therefore represents a serious threat to human health. In the present study, the mechanism of rabdoternin E against the proliferation of the lung cancer cell line A549 was explored. It was found that rabdoternin E caused the accumulation of large amounts of reactive oxygen species (ROS), promoted cell S phase arrest by reducing the expression of CDK2 and cyclin A2, induced apoptosis by increasing the Bax/Bcl‑2 ratio and promoted the phosphorylation of proteins in the ROS/p38 MAPK/JNK signaling pathway, which is associated with apoptosis and ferroptosis. In addition, it was also found that Z‑VAD‑FMK (an apoptosis inhibitor), ferrostatin‑1 (ferroptosis inhibitor) and N‑acetylcysteine (a ROS inhibitor) could partially or greatly reverse the cytotoxicity of rabdoternin E to A549 cells. Similarly, NAC (N‑acetylcysteine) treatment notably inhibited the rabdoternin E‑stimulated p38 MAPK and JNK activation. Furthermore, in vivo experiments in mice revealed that Rabdoternin E markedly reduced tumor volume and weight and regulated the expression levels of apoptosis and ferroptosis‑related proteins (including Ki67, Bcl‑2, Bax, glutathione peroxidase 4, solute carrier family 7 member 11 and transferrin) in the tumor tissues of mice. Histopathological observation confirmed that the number of tumor cells decreased markedly after administration of rabdoternin E. Taken together, rabdoternin E induced apoptosis and ferroptosis of A549 cells by activating the ROS/p38 MAPK/JNK signaling pathway. Therefore, the results of the present study showed that rabdoternin E is not toxic to MCF‑7 cells (normal lung cells), had no significant effect on body weight and was effective and therefore may be a novel therapeutic treatment for lung cancer.