Unveiling the unexplored novel signatures for osteoporosis via a detailed bioinformatics and molecular experiments based approach.

BACKGROUND: Osteoporosis (OP) stands as a prevalent bone ailment affecting the elderly, globally. The identification of reliable diagnostic markers crucially aids OP clinical management. METHODS: Utilizing the GEO database (GSE35959), we acquired expression profiles for OP and normal samples. Differential expression genes (DEGs) and hub genes were pinpointed through STRING, GEO2R, and Cytoscape. The competing endogenous RNA (ceRNA) network was constructed using miRTarBase, miRDB, and MiRcode databases. Gene Ontology (GO) and KEGG pathway enrichment analyses were performed via DAVID. Validation involved clinical OP samples from the Pakistani population, with Real-Time Quantitative Polymerase Chain Reaction (RT-qPCR) assessing hub gene expression. RESULTS: A total of 2124 differentially expressed genes (DEGs) were identified between OP and normal samples in GSE35959. The selected hub genes among these DEGs were Splicing Factor 3a Subunit 1 (SF3A1), Ataxin 2 Like (ATXN2L), Heat Shock Protein 90 Beta Family Member 1 (HSP90B1), Cluster of Differentiation 74 (CD74), DExH-Box Helicase 29 (DHX29), ALG5 Dolichyl-Phosphate Beta-Glucosyltransferase (ALG5), NudC Domain Containing 2 (NUDCD2), and Ras-related protein Rab-2A (RAB2A). Expression validation of these genes on the Pakistani OP patients revealed significant up-regulation of SF3A1, ATXN2L, and CD74 and significant (P < 0.05) down-regulation of HSP90B1, DHX29, ALG5, NUDCD2, and RAB2A in OP patients. Receiver operating characteristic (ROC) analysis demonstrated that these hub genes displayed considerable diagnostic accuracy for detecting OP. The ceRNA network analysis of the hub genes revealed some important hub genes' regulatory miRNAs and lncRNAs. Via KEGG analysis, hub genes were found to be enriched in N-Glycan biosynthesis, Thyroid hormone synthesis, IL-17 signaling pathway, Prostate cancer, AMPK signaling pathway, Spliceosome, Estrogen signaling pathway, and Fluid shear stress and atherosclerosis, etc., pathways. CONCLUSION: The identified eight hub genes in the present study could reliably distinguish OP patients from normal individuals, which may provide novel insight into the diagnostic research of OP.

Transcriptomics data integration and analysis to uncover hallmark genes in hypertrophic cardiomyopathy.

INTRODUCTION: Hypertrophic cardiomyopathy (HCM) is a heterogeneous disease that mainly affects the myocardium. In the current study, we aim to explore HCM-related hub genes through the analysis of differentially expressed genes (DEGs) between HCM and normal sample groups. METHODS: The GSE68316 and GSE36961 expression profiles were obtained from the Gene Expression Omnibus (GEO) database for the identification of DEGs, to explore hub genes, and to perform their expression analysis. Clinical HCM and control tissue samples were taken for expression and promoter methylation validation analysis via RNA-sequencing (RNA-seq) and targeted bisulfite sequencing (bisulfite-seq) analyses. Then, other different bioinformatics tools were employed to perform STRING, lncRNA-miRNA-mRNA regulatory networks, gene enrichment, and drug prediction analyses. RESULTS: In total, the top 20 DEGs, including 10 up-regulated and 10 down-regulated, were obtained from GSE68316. Out of the 20 DEGs, we subsequently identified the 8 most important hub genes including 5 up-regulated genes (EPB42, UQCRH, CA1, PFDN5, and LSM5) and 3 down-regulated genes (RPS24, TNS1, and RPL26). Expression and promoter methylation dysregulation of these genes were further validated on clinical HCM samples paired with controls. Next, we further investigated hub genes' regulatory 6 miRNAs (has-mir-1-3p, has-mir-129-5p, has-mir-16-5p, has-mir-23b-3p, has-mir-27-3p, and has-mir-182-5p) and miRNAs regulatory 4 lncRNAs (NUTMB2-AS1, NEAT1, XIST, and GABPB1-AS1) in this study via the lncRNA-cricRNA-miRNA-mRNA regulatory network. Later on, gene enrichment analysis revealed that hub genes were enriched in various important pathways including Nitrogen metabolism, Ribosome, RNA degradation, Cardiac muscle contraction, and Coronavirus disease, etc. Finally, the drug prediction analysis highlighted different potential candidate drugs for altering the expression of hub genes in the treatment of HCM. CONCLUSION: In summary, the identification of key hub genes and their enrichment analysis in the current study may shed light on the mechanisms behind the occurrence and development of HCM.

Structural Dynamics of P-Rex1 Complexed with Natural Leads Establishes the Protein as an Attractive Target for Therapeutics to Suppress Cancer Metastasis.

Phosphatidylinositol 3,4,5-trisphosphate- (PIP3-) dependent Rac exchanger 1 (P-Rex1) functions as Rho guanine nucleotide exchange factor and is activated by synergistic activity of Gßγ and PIP3 of the heterotrimeric G protein. P-Rex1 activates Rac GTPases for regulating cell invasion and migration and promotes metastasis in several human cancers including breast, prostate, and skin cancer. The protein is a promising therapeutic target because of its multifunction roles in human cancers. Herein, the present study attempts to identify selective P-Rex1 natural inhibitors by targeting PIP3-binding pocket using large-size multiple natural molecule libraries. Each library was filtered subsequently in FAF-Drugs4 based on Lipinski's rule of five (RO5), toxicity, and filter pan assay interference compounds (PAINS). The output hits were virtually screened at the PIP3-binding pocket through PyRx AutoDock Vina and cross-checked by GOLD. The best binders at the PIP3-binding pocket were prioritized using a comparative analysis of the docking scores. Top-ranked two compounds with high GOLD fitness score (>80) and lowest AutoDock binding energy (< -12.7 kcal/mol) were complexed and deciphered for molecular dynamics along with control-P-Rex1 complex to validate compound binding conformation and disclosed binding interaction pattern. Both the systems were seen in good equilibrium, and along the simulation time, the compounds are in strong contact with the P-Rex1 PIP3-binding site. Hydrogen bonding analysis towards simulation end identified the formation of 16 and 22 short- and long-distance hydrogen bonds with different percent of occupancy to the PIP3 residues for compound I and compound 2, respectively. Radial distribution function (RDF) analysis of the key hydrogen bonds between the compound and the PIP3 residues demonstrated a strong affinity of the compounds to the mentioned PIP3 pocket. Additionally, MMGB/PBSA energies were performed that confirmed the dominance of Van der Waals energy in complex formation along with favorable contribution from hydrogen bonding. These findings were also cross-validated by a more robust WaterSwap binding energy predictor, and the results are in good agreement with a strong binding affinity of the compounds for the protein. Lastly, the key contribution of residues in interaction with the compounds was understood by binding free energy decomposition and alanine scanning methods. In short, the results of this study suggest that P-Rex1 is a good druggable target to suppress cancer metastasis; therefore, the screened druglike molecules of this study need in vitro and in vivo anti-P-Rex1 validation and may serve as potent leads to fight cancer.

Assessment of Two Potential Probiotic Strains As Anti-Obesity Supplements Under High-Fat Feeding Conditions.

Obesity is one of the chronic diseases that increase annually and cause cardiovascular disease, which is the main cause of death worldwide. So, this study aims to evaluate the role of the two potential probiotics: Lactiplantibacillus plantarum Pro1 and Lacticaseibacillus rhamnosus Pro2, isolated from the fermented milk and corn silage as anti-obesity supplements. Seventy-five male BALB/c mice were distributed to five groups (control, obesity, obesity plus L. plantarum (OLP), obesity plus L. rhamnosus (OLR) and obesity plus mixture of two strains (OM)). The body weight, lipid profile, histopathology and enzymes of liver were assessed. RT-PCR was used to determine the expression of CYP7A1, ALTG4, TNFα and ROR genes.The findings show that the obesity group recorded the significant highest value of the body weight, TC, TG, LDL, AST and ALT, while OLP group recorded the significant lowest value. Liver tissue of obesity group has necrosis and fatty changes, while the OLP group was related to the control group. The findings of RT-PCR show non-significant differences between the control group and the OLP group, with significant differences between the control group and the set groups in expression of CYP7A1, ALTG4, TNFα and ROR genes. L. plantarum Pro1 reduced the expression of inflammation genes (TNFα and ROR), and increase the expression of the lipid metabolism genes (CYP7A1, ALTG4) to reduce the inflammatory effects of obesity in the liver, and decrease the cholesterol level in serum. Therefore, L. plantarum Pro1 is useful as anti-obesity supplements and an eliminator of the relevant diseases.

Synthesis, Characterization, and Antimicrobial and Antiproliferative Effects of CuO-TiO2-Chitosan-Escin Nanocomposites on Human Leukemic MOLT4 Cells.

Nanocomposites comprised of CuO-TiO2-chitosan-escin, which has adjustable physicochemical properties, provide a solution for therapeutic selectivity in cancer treatment. By controlling the intrinsic signaling primarily through the mitochondrial signaling pathway, we desired nanocomposites with enhanced anticancer activity by containing CuO-TiO2-chitosan-escin. The metal oxides CuO and TiO2, the natural polymer chitosan, and a phytochemical compound escin were combined to form CuO-TiO2-chitosan-escin nanocomposites. The synthesized nanocomposites were confirmed and characterized using FTIR spectroscopy, TEM, and UV-Vis absorption spectroscopy. A human leukemia cell line (MOLT-4) was used to assess the efficacy and selectivity of nanocomposites. Based on a cytotoxicity study, CuO-TiO2-chitosan-escin nanocomposites had inhibition concentrations (IC50) of 13.68, 8.9, and 7.14 µg/mL against human T lymphoblast cells after 24, 48, and 72 h of incubation, respectively. Compared with untreated MOLT-4 cells, CuO-TiO2-chitosan-escin nanocomposite-treated cells significantly increased (p < 0.05) caspase-3, -8, and -9 and decreased the levels of antioxidant enzymes GR, SOD, and GSH. Furthermore, MDA for lipid peroxidase and ROS levels significantly increased (p < 0.05) in the treated cells than in the untreated cells. Remarkably, CuO-TiO2-chitosan-escin nanocomposite-mediated control of cell cycles were mainly achieved through the activation of caspase-3, -8, and -9.

Could allicin alleviate trastuzumab-induced cardiotoxicity in a rat model through antioxidant, anti-inflammatory, and antihyperlipidemic properties?

AIMS: Although trastuzumab (TZB)-induced cardiotoxicity is well documented and allicin (one of the main active garlic ingredients) has ameliorating effects against numerous causes of toxicities; however, the influence of allicin on TZB-induced cardiotoxicity has not been investigated yet. Therefore, the current work explored the potential cardioprotective structural, biochemical, and molecular mechanisms of allicin against TZB-induced cardiotoxicity in a rat's model. METHODS: Forty rats were divided into four equal groups and treated for five weeks. The control group (G1) received PBS, the allicin group (G2) received allicin (9 mg/kg/day), the TZB group (G3) received TZB (6 mg/kg/week), and the allicin+TZB group (G4) received 9 mg of allicin/kg/day +6 mg of TZB/kg/week. Heart specimens and blood samples were processed for histopathological, immunohistochemical, biochemical, and molecular investigations to determine the extent of cardiac injury in all groups. KEY FINDINGS: The myocardium of G3 revealed significant increases in the numbers of inflammatory and apoptotic cells and the area percentage of collagen fibers and TNF-α immunoexpression compared with G1 and G2. Besides, qRT-PCR analysis exhibited significant reductions of SOD3, GPX1, and CAT expressions with significant increases in TNFα, IL-1ß, IL-6, cTnI, cTnT, and LDH expressions. Additionally, flow cytometry analysis demonstrated a significant elevation in the apoptotic and ROS levels. In contrast, allicin+TZB cotherapy in G4 ameliorated all previous changes compared with G3. SIGNIFICANCE: The current study proves that allicin could be used as a novel supplementary cardioprotective therapy to avoid TZB-induced cardiotoxicity via its anti-inflammatory, antifibrotic, antioxidant, antihyperlipidemic, and antiapoptotic properties.

Experimental and Theoretical Insights on Chemopreventive Effect of the Liposomal Thymoquinone Against Benzo[a]pyrene-Induced Lung Cancer in Swiss Albino Mice.

Purpose: Thymoquinone (TQ), a phytoconstituent of Nigella sativa seeds, has been studied extensively in various cancer models. However, TQ's limited water solubility restricts its therapeutic applicability. Our work aims to prepare the novel formulation of TQ and assess its chemopreventive potential in chemically induced lung cancer animal model. Methods: The polyethylene glycol coated DOPE/CHEMS incorporating TQ-loaded pH-sensitive liposomes (TQPSL) were prepared and characterized. Mice were exposed to benzo[a]pyrene (BaP) thrice a week for 4 weeks to induce lung cancer. TQPSL was administered three times a week for 21 weeks, starting 2 weeks before the first dose of BaP. Results: The prepared TQPSL revealed 85% entrapment efficiency with 128 nm size and -19.5 mv ζ-potential showing high stability of the formulation. The pretreatment of TQPSL showed the recovery in BaP-modulated relative organ weight of lungs, cancer marker enzymes, and antioxidant enzymes in the serum. The histopathological analysis of the tissues showed that TQPSL protected the malignancy in the lungs. The flow cytometry data revealed the induction of apoptosis and decreased intracellular ROS by TQPSL. Molecular docking was performed to predict the TQ's affinity for eight possible anticancer drug targets linked to lung cancer etiology. The data assisted to identify the serine/threonine-protein kinase BRAF as the most suitable target of TQ with binding energy -6.8 kcal/mol. Conclusion: The current findings demonstrated the potential of TQPSL and its possible therapeutic targets of lung cancer. To our knowledge, this is the first research to outline the development of TQ formulation against lung cancer considering its low solubility as well as pulmonary delivery challenges.

Cytoprotective Antioxidant, Anti-Inflammatory, and Antifibrotic Impact of Celery Seed Oil and Manuka Honey Against Cyclophosphamide-Induced Cystitis in Rabbits.

Patients treated with cyclophosphamide (CP) usually suffer from severe hemorrhagic cystitis (HC). Our previous study exhibited that mesna + celery cotherapy partially ameliorated HC. Therefore, there is a substantial need to seek alternative regimens to get complete protection against CP-induced HC. The current study investigated the effects of mesna + celery seed oil (MCSO) or mesna + manuka honey (MMH) cotherapy against CP-induced HC in adult male rabbits. The forty rabbits were divided into four equal groups and treated for three weeks. The control group (G1) received distilled water and the second group (G2) received CP (50 mg/kg/week). The third group (G3) received CP + MCSO (CPMCSO regimen), and the fourth group (G4) received CP + MMH (CPMMH regimen). The urinary bladder (UB) specimens were processed to evaluate UB changes through histopathological, immunohistochemical, ultrastructural, and biochemical investigations. In G2, CP provoked HC features (urothelial necrosis, ulceration, and sloughing), UB fibrosis, and TNF-α immunoexpression. Besides, CP reduced the activity of antioxidant enzymes (GPx1, SOD3, and CAT) and elevated the serum levels of NF-κB, TNF-α, IL-1B, and IL-6 cytokines in G2 rabbits. In contrast, the CPMMH regimen caused significant increments of UB protection against HC in G4 rabbits compared to the partial protection by the CPMCSO regimen in G3. Therefore, our study indicated for the first time that the novel CPMMH regimen resulted in complete UB protection against CP-induced HC via combined antioxidant, anti-inflammatory, and antifibrotic properties.

The Effect of Liposomal Diallyl Disulfide and Oxaliplatin on Proliferation of Colorectal Cancer Cells: In Vitro and In Silico Analysis.

Diallyl disulfide (DADS) is one of the main bioactive organosulfur compounds of garlic, and its potential against various cancer models has been demonstrated. The poor solubility of DADS in aqueous solutions limits its uses in clinical application. The present study aimed to develop a novel formulation of DADS to increase its bioavailability and therapeutic potential and evaluate its role in combination with oxaliplatin (OXA) in the colorectal cancer system. We prepared and characterized PEGylated, DADS (DCPDD), and OXA (DCPDO) liposomes. The anticancer potential of these formulations was then evaluated in HCT116 and RKO colon cancer cells by different cellular assays. Further, a molecular docking-based computational analysis was conducted to determine the probable binding interactions of DADS and OXA. The results revealed the size of the DCPDD and DCPDO to be 114.46 nm (95% EE) and 149.45 nm (54% EE), respectively. They increased the sensitivity of the cells and reduced the IC50 several folds, while the combinations of them showed a synergistic effect and induced apoptosis by 55% in the cells. The molecular docking data projected several possible targets of DADS and OXA that could be evaluated more precisely by these novel formulations in detail. This study will direct the usage of DCPDD to augment the therapeutic potential of DCPDO against colon cancer in clinical settings.

Neuroprotective Effect of Chrysophanol as a PI3K/AKT/mTOR Signaling Inhibitor in an Experimental Model of Autologous Blood-induced Intracerebral Hemorrhage.

OBJECTIVE: Intracerebral hemorrhage (ICH) refers to predominant, sporadic, and non-traumatic bleeding in the brain parenchyma. The PI3K/AKT/mTOR signaling pathway is an important signal transduction pathway regulated by enzyme-linked receptors and has many biological functions in mammals. It plays a key role in neuronal metabolism, gene expression regulation, and tissue homeostasis in the healthy and diseased brain. METHODS: In the present study, the role of the PI3K/AKT/mTOR pathway inhibitor chrysophanol (CPH) (10 mg/kg and 20 mg/kg, orally) in the improvement of ICH-associated neurological defects in rats was investigated. Autologous blood (20 µL/5 min/unilateral/intracerebroventricular) mimics ICH-like defects involving cellular and molecular dysfunction and neurotransmitter imbalance. The current study also included various behavioral assessments to examine cognition, memory, and motor and neuromuscular coordination. The protein expression levels of PI3K, AKT, and mTOR as well as myelin basic protein and apoptotic markers, such as Bax, Bcl-2, and caspase-3, were examined using ELISA kits. Furthermore, the levels of various neuroinflammatory cytokines and oxidative stress markers were assessed. Additionally, the neurological severity score, brain water content, gross brain pathology, and hematoma size were used to indicate neurological function and brain edema. RESULTS: CPH was found to be neuroprotective by restoring neurobehavioral alterations and significantly reducing the elevated PI3K, AKT, and mTOR protein levels, and modulating the apoptotic markers such as Bax, Bcl-2, and caspase-3 in rat brain homogenate. CPH substantially reduced the inflammatory cytokines like interleukin (IL)-1ß, IL-6, and tumor necrosis factor-α. CPH administration restored the neurotransmitters GABA, glutamate, acetylcholine, dopamine, and various oxidative stress markers. CONCLUSION: Our results show that CPH may be a promising therapeutic approach for overcoming neuronal damage caused by the overexpression of the PI3K/AKT/mTOR signaling pathway in ICH-induced neurological dysfunctions in rats.

Effect of obesity on fertility parameters in WIO mice model.

BACKGROUND: Male infertility is mostly due to low sperm quality, which accounts for about 50% of the causes of infertility. The reasons for low sperm quality are still unclear. Nowadays, many drinks contain high levels of fat, and its effect on fertility is not yet known. OBJECTIVES: To investigate the effect of cholesterol-containing water on male fertility. MATERIAL AND METHODS: Forty BALB/c male mice were divided into 2 groups: the control group and the water-induced obesity (WIO) group. Body and testicular weights were recorded and analyzed statistically. Testicular tissues were examined. Serum contents of total cholesterol (TC), triglycerides (TG), low-density lipoprotein (LDL), free testosterone (FT), luteinizing hormone (LH), and follicle-stimulating hormone (FSH) were determined. Motility count and morphology of sperm were analyzed. Real-time polymerase chain reaction (RT-PCR) was performed for SYCP3, VEGFA and WT1 genes. RESULTS: The results showed that the WIO group presented the highly significant values for mice body and testis weight, and TC, TG and LDL level in serum (p < 0.05), when compared to the control group. The level of FT, LH and FSH in serum was significantly decreased (p < 0.05) in the WIO group compared with the control group. Seminiferous tubules of testes became thin, and Sertoli cells showed mild atrophy in this group. Also, the count and motility of sperm significantly reduced while the ratio of sperm abnormalities significantly increased in the WIO group compared with the control group (p < 0.05). The results of RT-PCR showed that SYCP3, VEGFA and WT1 genes were significantly downregulated (p < 0.05) in the WIO group compared with the control group. CONCLUSIONS: This study indicated that drinks containing high levels of fat may have negative effects on male fertility due to the reduction of the sexual hormones level in serum, the expression of SYCP3, VEGFA and WT1 genes, count and motility of sperm, as well as an increase in sperm abnormalities and pathological changes in the testicular tissues.

Neuroprotective Effect of Chrysophanol as a PI3K/AKT/mTOR Signaling Inhibitor in an Experimental Model of Autologous Blood-induced Intracerebral Hemorrhage.

OBJECTIVE: Intracerebral hemorrhage (ICH) refers to predominant, sporadic, and non-traumatic bleeding in the brain parenchyma. The PI3K/AKT/mTOR signaling pathway is an important signal transduction pathway regulated by enzyme-linked receptors and has many biological functions in mammals. It plays a key role in neuronal metabolism, gene expression regulation, and tissue homeostasis in the healthy and diseased brain. METHODS: In the present study, the role of the PI3K/AKT/mTOR pathway inhibitor chrysophanol (CPH) (10 mg/kg and 20 mg/kg, orally) in the improvement of ICH-associated neurological defects in rats was investigated. Autologous blood (20 µL/5 min/unilateral/intracerebroventricular) mimics ICH-like defects involving cellular and molecular dysfunction and neurotransmitter imbalance. The current study also included various behavioral assessments to examine cognition, memory, and motor and neuromuscular coordination. The protein expression levels of PI3K, AKT, and mTOR as well as myelin basic protein and apoptotic markers, such as Bax, Bcl-2, and caspase-3, were examined using ELISA kits. Furthermore, the levels of various neuroinflammatory cytokines and oxidative stress markers were assessed. Additionally, the neurological severity score, brain water content, gross brain pathology, and hematoma size were used to indicate neurological function and brain edema. RESULTS: CPH was found to be neuroprotective by restoring neurobehavioral alterations and significantly reducing the elevated PI3K, AKT, and mTOR protein levels, and modulating the apoptotic markers such as Bax, Bcl-2, and caspase-3 in rat brain homogenate. CPH substantially reduced the inflammatory cytokines like interleukin (IL)-1ß, IL-6, and tumor necrosis factor-α. CPH administration restored the neurotransmitters GABA, glutamate, acetylcholine, dopamine, and various oxidative stress markers. CONCLUSION: Our results show that CPH may be a promising therapeutic approach for overcoming neuronal damage caused by the overexpression of the PI3K/AKT/mTOR signaling pathway in ICH-induced neurological dysfunctions in rats.

Towards A Novel Multi-Epitopes Chimeric Vaccine for Simulating Strong Immune Responses and Protection against Morganella morganii.

Morganella morganii is one of the main etiological agents of hospital-acquired infections and no licensed vaccine is available against the pathogen. Herein, we designed a multi-epitope-based vaccine against M. morganii. Predicted proteins from fully sequenced genomes of the pathogen were subjected to a core sequences analysis, followed by the prioritization of non-redundant, host non-homologous and extracellular, outer membrane and periplasmic membrane virulent proteins as vaccine targets. Five proteins (TonB-dependent siderophore receptor, serralysin family metalloprotease, type 1 fimbrial protein, flagellar hook protein (FlgE), and pilus periplasmic chaperone) were shortlisted for the epitope prediction. The predicted epitopes were checked for antigenicity, toxicity, solubility, and binding affinity with the DRB*0101 allele. The selected epitopes were linked with each other through GPGPG linkers and were joined with the cholera toxin B subunit (CTBS) to boost immune responses. The tertiary structure of the vaccine was modeled and blindly docked with MHC-I, MHC-II, and Toll-like receptors 4 (TLR4). Molecular dynamic simulations of 250 nanoseconds affirmed that the designed vaccine showed stable conformation with the receptors. Further, intermolecular binding free energies demonstrated the domination of both the van der Waals and electrostatic energies. Overall, the results of the current study might help experimentalists to develop a novel vaccine against M. morganii.

Protective and therapeutic effects of two novel strains of Lactobacilli on diabetes-associated disorders induced by a high level of fructose.

Diabetes is a metabolic disorder described as insufficient secretion of insulin in the pancreas or the inability of the existing insulin to function properly. It poses a greater risk on human health as it is considered the base of several diseases. Thus, this study was designed to evaluate two novel strains of Lactobacillus in handling pancreas disorders. 50 BALB/c male mice were divided into five groups; (a) feeding on normal diet only as control group, (b) given 21% fructose in drinking water as diabetes group, (c) feeding on Lactobacillus rhamnosus strain Pro2 (MT505335.1) plus 21% fructose as LR group, (d) feeding on Lactobacillus plantarum strain Pro1 (MT505334.1) plus 21% fructose as LP group and (e) mixture of two strains plus 21% fructose as Mix group. The serum content of glucose, aspartate aminotransferase (AST), and alanine aminotransferase (ALT) was determined. Pancreases histopathology was examined. Expression of GH, IGF1, and GLP-1 genes was measured in the liver and pancreas by RT-qPCR. Serum content of glucose, ALT, and AST significantly increased in diabetes group, and significantly reduced in (LP) and (Mix) groups compared with control. Pathological changes occurred in the exocrine and endocrine components of the diabetes group pancreas. Besides, islet cells are almost entirely disturbed and acinar cells degenerated. However, in (LP) and (Mix) groups, the pathological changes significantly decreased and became related to the control group. Expression of GH, IGF1, and GLP-1 genes was significantly downregulated in the liver and pancreas of mice given fructose compared with control. Expression of these genes was either significantly upregulated in groups (LP and Mix) or identical to the control group. This study shows that the strain Pro1 (MT505334.1) or a combination of two strains is useful in reducing diabetic risk.

Central Administration of Insulin Combined With Resistin Reduces Renal Sympathetic Nerve Activity in Rats Fed a High Fat Diet.

Insulin receptors are widely distributed in the central nervous system and their activation by insulin elicits renal sympatho-excitatory effects. Resistin, an adipokine, promotes resistance to the metabolic effects of insulin. Resistin also induces increases in renal sympathetic nerve activity (RSNA) by acting in the brain, but whether it can influence insulin's actions on RSNA is unknown. In the present study we investigated, in male Sprague-Dawley rats (7-8 weeks of age), the effects of central administration of insulin combined with resistin on RSNA following a normal diet (ND) and a high fat diet (HFD) (22% fat), since HFD can reportedly attenuate insulin's actions. RSNA, mean arterial pressure (MAP) and heart rate (HR) responses were monitored and recorded before and for 180 min after intracerebroventricular injection of saline (control) (n = 5 HFD and ND), resistin (7 µg; n = 4 ND, n = 5 HFD), insulin (500 mU; n = 6 ND, n = 5 HFD), and the combination of both resistin and insulin (n = 7 ND, n = 5 HFD). The key finding of the present study was that when resistin and insulin were combined there was no increase in RSNA induced in rats fed a normal diet or the high fat diet. This contrasted with the sympatho-excitatory RSNA effects of the hormones when each was administered alone in rats fed the ND and the HFD.

High Fat Diet Decreases Neuronal Activation in the Brain Induced by Resistin and Leptin.

Resistin and leptin are adipokines which act in the brain to regulate metabolic and cardiovascular functions which in some instances are similar, suggesting activation of some common brain pathways. High-fat feeding can reduce the number of activated neurons observed following the central administration of leptin in animals, but the effects on resistin are unknown. The present work compared the distribution of neurons in the brain that are activated by centrally administered resistin, or leptin alone, and, in combination, in rats fed a high fat (HFD) compared to a normal chow diet (ND). Immunohistochemistry for the protein, Fos, was used as a marker of activated neurons. The key findings are (i) following resistin or leptin, either alone or combined, in rats fed the HFD, there were no significant increases in the number of activated neurons in the paraventricular and arcuate nuclei, and in the lateral hypothalamic area (LHA). This contrasted with observations in rats fed a normal chow diet; (ii) in the OVLT and MnPO of HFD rats there were significantly less activated neurons compared to ND following the combined administration of resistin and leptin; (iii) In the PAG, RVMM, and NTS of HFD rats there were significantly less activated neurons compared to ND following resistin. The results suggest that the sensitivity to resistin in the brain was reduced in rats fed a HFD. This has similarities with leptin but there were instances where there was reduced sensitivity to resistin with no significant effects following leptin. This suggests diet influences neuronal effects of resistin.

Central leptin and resistin combined elicit enhanced central effects on renal sympathetic nerve activity.

NEW FINDINGS: What is the central question of this study? Leptin and resistin act centrally to increase renal sympathetic nerve activity (RSNA). We investigated whether a combination of resistin and leptin could induce a greater response than either alone. We also used Fos protein to quantify the number of activated neurons in the brain. What is the main finding and its importance? A combination of leptin and resistin induced a greater increase in RSNA than either hormone alone. This was correlated with a greater number of activated neurons in the arcuate nucleus than with either hormone alone. Leptin and resistin act centrally to increase renal sympathetic nerve activity (RSNA). We investigated whether a combination of resistin and leptin could induce a greater response than either alone. Mean arterial pressure, heart rate and RSNA were recorded before and for 3 h after intracerebroventricular saline (control; n = 5), leptin (7 µg; n = 5), resistin (7 µg; n = 4) and leptin administered 15 min after resistin (n = 6). Leptin alone and resistin alone significantly increased RSNA (74 ± 17 and 50 ± 14%, respectively; P < 0.0001 compared with saline). When leptin and resistin were combined, there was a significantly greater increase in RSNA (163 ± 23%) compared with either hormone alone (P < 0.0001). Maximal responses of mean arterial pressure and heart rate were not significantly different between groups. We also used Fos protein to quantify the number of activated neurons in the brain. Compared with controls, there were significant increases in numbers of Fos-positive neurons in the arcuate and hypothalamic paraventricular nuclei when leptin or resistin was administered alone or when they were combined, and in the lamina terminalis when leptin and resistin were combined. Only in the arcuate nucleus was the increase significantly greater compared with either hormone alone. The findings show that a combination of leptin and resistin induces a greater RSNA increase and a greater number of activated neurons in the arcuate nucleus than with either hormone alone. Given that leptin makes an important contribution to the elevated RSNA observed in obese and overweight conditions, the increased concentrations of leptin and resistin may mean that the contribution of leptin to the elevated RSNA in those conditions is enhanced.

Central Administration of Insulin and Leptin Together Enhance Renal Sympathetic Nerve Activity and Fos Production in the Arcuate Nucleus.

There is considerable interest in the central actions of insulin and leptin. Both induce sympatho-excitation. This study (i) investigated whether centrally administered leptin and insulin together elicits greater increases in renal sympathetic nerve activity (RSNA), mean arterial pressure (MAP) and heart rate (HR) than when given alone, and (ii) quantified the number of activated neurons in brain regions influencing SNA, to identify potential central sites of interaction. In anesthetised (urethane 1.4-1.6 g/kg iv) male Sprague-Dawley rats, RSNA, MAP, and HR were recorded following intracerebroventricular (ICV) saline (control; n = 5), leptin (7 µg; n = 5), insulin (500 mU; n = 4) and the combination of leptin and insulin; (n = 4). Following leptin or insulin alone, RSNA was significantly increased (74 and 62% respectively). MAP responses were not significantly different between the groups. Insulin alone significantly increased HR. Leptin alone also increased HR but it was significantly less than following insulin alone (P < 0.005). When leptin and insulin were combined, the RSNA increase (124%) was significantly greater than the response to either alone. There were no differences between the groups in MAP responses, however, the increase in HR induced by insulin was attenuated by leptin. Of the brain regions examined, only in the arcuate nucleus did leptin and insulin together increase the number of Fos-positive cell nuclei significantly more than leptin or insulin alone. In the lamina terminalis and rostroventrolateral medulla, leptin and insulin together increased Fos, but the effect was not greater than leptin alone. The results suggest that when central leptin and insulin levels are elevated, the sympatho-excitatory response in RSNA will be greater. The arcuate nucleus may be a common site of cardiovascular integration.