This study aimed to explore the mechanisms and molecular targets of total flavones of Abelmoschus manihot(TFA) plus empagliflozin(EM) in attenuating diabetic tubulopathy(DT) by targeting mitochondrial homeostasis and pyroptosis-apoptosis-necroptosis(PANoptosis). In the in vivo study, the authors established the DT rat models through a combination of uninephrectomy, administration of streptozotocin via intraperitoneal injections, and exposure to a high-fat diet. Following modeling successfully, the DT rat models received either TFA, EM, TFA+EM, or saline(as a vehicle) by gavage for eight weeks, respectively. In the in vitro study, the authors subjected the NRK52E cells with or without knock-down Z-DNA binding protein 1(ZBP1) to a high-glucose(HG) environment and various treatments including TFA, EM, and TFA+EM. In the in vivo and in vitro studies, The authors investigated the relative characteristics of renal tubular injury and renal tubular epithelial cells damage induced by reactive oxygen species(ROS), analyzed the relative characteristics of renal tubular PANoptosis and ZBP1-mediatted PANoptosis in renal tubular epithelial cells, and compared the relative characteristics of the protein expression levels of marked molecules of mitochondrial fission in the kidneys and mitochondrial homeostasis in renal tubular epithelial cells, respectively. Furthermore, in the network pharmacology study, the authors predicted and screened targets of TFA and EM using HERB and SwissTargetPrediction databases; The screened chemical constituents and targets of TFA and EM were constructed the relative network using Cytoscape 3.7.2 network graphics software; The relative targets of DT were integrated using OMIM and GeneCards databases; The intersecting targets of TFA, EM, and DT were enriched and analyzed signaling pathways by Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG) software using DAVID database. In vivo study results showed that TFA+EM could improve renal tubular injury, the protein expression levels and characteristics of key signaling molecules in PANoptosis pathway in the kidneys, and the protein expression levels of marked molecules of mitochondrial fission in the kidneys. And that, the ameliorative effects in vivo of TFA+EM were both superior to TFA or EM. Network pharmacology study results showed that TFA+EM treated DT by regulating the PANoptosis signaling pathway. In vitro study results showed that TFA+EM could improve ROS-induced cell injury, ZBP1-mediatted PANoptosis, and mitochondrial homeostasis in renal tubular epithelial cells under a state of HG, including the protein expression levels of marked molecules of mitochondrial fission, mitochondrial ultrastructure, and membrane potential level. And that, the ameliorative effects in vitro of TFA+EM were both superior to TFA or EM. More importantly, using the NRK52E cells with knock-down ZBP1, the authors found that, indeed, ZBP1 was mediated PANoptosis in renal tubular epithelial cells as an upstream factor. In addition, TFA+EM could regulate the protein expression levels of marked signaling molecules of PANoptosis by targeting ZBP1. In summary, this study clarified that TFA+EM, different from TFA or EM, could attenuate DT with multiple targets by ameliorating mitochondrial homeostasis and inhibiting ZBP1-mediated PANoptosis. These findings provide the clear pharmacological evidence for the clinical treatment of DT with a novel strategy of TFA+EM, which is named "coordinated traditional Chinese and western medicine".
Animals ; Rats ; Mitochondria/metabolism* ; Benzhydryl Compounds/administration & dosage* ; Glucosides/administration & dosage* ; Abelmoschus/chemistry* ; Male ; Homeostasis/drug effects* ; Flavones/administration & dosage* ; Rats, Sprague-Dawley ; Diabetic Nephropathies/physiopathology* ; Drugs, Chinese Herbal/administration & dosage* ; DNA-Binding Proteins/genetics* ; Humans ; Apoptosis/drug effects*

OBJECTIVETo observe the influence of different concentrations of bisphenol A (BPA) on glucose metabolism and lactate dehydrogenase (LDH) expression in rat Sertoli cells in vitro and investigate the mechanisms of BPA inducing male infertility.

METHODSUsing two-step enzyme digestion, we isolated Sertoli cells from male Wistar rats and constructed a primary Sertoli cell system, followed by immunohistochemical FasL staining. We randomly divided the Sertoli cells into a control group to be cultured in the serum-free minimal essential medium (MEM) plus dimethyl sulfoxide (DMSO) and three experimental groups to be treated with 100 nmol/L, 10 μmol/L, and 1 mmol/L BPA, respectively, in the MEM plus DMSO. After 48 hours of treatment, we measured the proliferation of the cells by CCK-8 assay, determined the concentrations of metabolites by NMR spectroscopy, and detected the expression of LDH in the Sertoli cells by RT-PCR and Western blot.

RESULTSThe purity of the isolated Sertoli cells was (96.05 ± 1.28)% (n = 10). Compared with the control group, the 100 nmol/L, 10 μmol/L, and 1 mmol/L BPA groups showed no remarkable changes in the proliferation of Sertoli cells ([98 ± 8]%, [96 ± 3]%, and [95 ± 3]%, P >0.05), but the 10 μmol/L and 1 mmol/L of BPA groups exhibited significantly decreased concentrations of intracellular glucose ([3.89 ± 0.07] vs [3.36 ± 0.24] and [3.04 ± 0.21] pmol/cell, P <0.05) and lactate ([0.43 ± 0.06] vs [0.29 ± 0.05] and [0.20 ± 0.03] pmol/cell, P <0.05). The expression of LDH mRNA was decreased with the increased concentration of BPA, while that of LDH protein reduced only in the 1 mmol/L BPA group (P <0.05).

CONCLUSIONHigh-concentration BPA decreases the expression of LDH and alters glucose metabolism in Sertoli cells, and therefore may reduce the provision of lactate for germ cells and impair spermatogenesis.

Animals ; Benzhydryl Compounds ; administration & dosage ; pharmacology ; Cell Proliferation ; drug effects ; Cells, Cultured ; Culture Media, Serum-Free ; Dimethyl Sulfoxide ; pharmacology ; Glucose ; metabolism ; In Vitro Techniques ; Infertility, Male ; chemically induced ; L-Lactate Dehydrogenase ; metabolism ; Male ; Phenols ; administration & dosage ; pharmacology ; RNA, Messenger ; metabolism ; Random Allocation ; Rats ; Rats, Wistar ; Sertoli Cells ; drug effects ; metabolism ; Spermatogenesis ; drug effects

OBJECTIVETo evaluate the combined subchronic toxicity of bisphenol A (BPA) and dibutyl phthalate (DBP) in male Sprague Dawley (SD) rats.

METHODSForty 4-week-old male rats weighing 115-125 g were randomly divided into BPA-treated, DBP-treated group, BPA+DBP-treated and control groups and fed with a soy- and alfalfa-free diet containing 285.4 ppm BPA, 285.4 ppm DBP, 285.4 ppm BPA plus 285.4 ppm DBP, and a control diet, respectively, for 90 consecutive days. At the end of the study, the animals were sacrificed by exsanguination via the carotid artery under diethyl etherane aesthesia and weighed. Organs, including liver, kidneys, spleen, thymus, heart, brain, and testis underwent pathological examination. The androgen receptor (AR), gonadotropin-releasing hormone receptor (GNRHR), and progesterone hormone receptor (PR) genes from the hypothalamus were detected by real-time PCR. The biomedical parameters were analyzed.

RESULTSNo significant difference was found in food intake, body weight, tissue weight, organ/brain weight ratio, and biomedical parameters among the four groups (P>0.05). However, BPA and DBP up-regulated AR, PR and GNRHR expression levels in rats and showed a synergistic or an additive effect in the BPA+DBP group.

CONCLUSIONThe combined subchronic toxicity of BPA and DBP is synergistic or additive in male SD rats.

Animals ; Benzhydryl Compounds ; administration & dosage ; toxicity ; Body Weight ; drug effects ; Dibutyl Phthalate ; administration & dosage ; toxicity ; Drug Interactions ; Eating ; drug effects ; Environmental Pollutants ; administration & dosage ; toxicity ; Male ; Phenols ; administration & dosage ; toxicity ; Random Allocation ; Rats ; Rats, Sprague-Dawley

To guide the reasonable clinical application of modafinil (MOD), pharmacokinetics and pharmacodynamics of MOD in mice and the correlation between them were investigated. Male mice (Kunming strain) were given a single oral dose of MOD (120 mg x kg(-1)). The plasma concentration of MOD was measured by HPLC and the pharmacokinetic parameters were calculated with DAS 3.0 software. For another batch of male Kunming strain mice, their locomotor activities were recorded by an infrared ray passive sensor after a same oral dose of MOD, and the synchronization and correlation between the changes of MOD plasma concentration and the locomotor activity induced by MOD were compared and analyzed. The results showed that the plasma concentration-time curve of MOD was fitted to two-compartment open model with a first order absorption. The main pharmacokinetic parameters t1/2alpha, t1/2beta, t(max), C(max) and AUC(0-inifinity) were 0.42 h, 3.10 h, 1.00 h, 41.34 mg x L(-1) and 142.22 mg x L(-1) x h, respectively. MOD significantly increased locomotor activity and the effect lasted for about 4 h. The changes of MOD plasma concentration and the locomotor activity induced by MOD were synchronous. In conclusion, there is a significant correlation between the effect of MOD and its plasma concentration after administration of 120 mg x kg(-1) in mice.
Administration, Oral ; Animals ; Area Under Curve ; Benzhydryl Compounds ; administration & dosage ; blood ; pharmacokinetics ; pharmacology ; Central Nervous System Stimulants ; administration & dosage ; blood ; pharmacokinetics ; pharmacology ; Chromatography, High Pressure Liquid ; Dose-Response Relationship, Drug ; Male ; Mice ; Motor Activity ; drug effects

The in vivo muscarinic receptor binding of antimuscarinic agents (oxybutynin, solifenacin, tolterodine, and imidafenacin) used to treat urinary dysfunction in patients with overactive bladder is reviewed. Transdermal administration of oxybutynin in rats leads to significant binding of muscarinic receptors in the bladder without long-term binding in the submaxillary gland and the abolishment of salivation evoked by oral oxybutynin. Oral solifenacin shows significant and long-lasting binding to muscarinic receptors in mouse tissues expressing the M3 subtype. Oral tolterodine binds more selectively to muscarinic receptors in the bladder than in the submaxillary gland in mice. The muscarinic receptor binding of oral imidafenacin in rats is more selective and longer-lasting in the bladder than in other tissues such as the submaxillary gland, heart, colon, lung, and brain, suggesting preferential muscarinic receptor binding in the bladder. In vivo quantitative autoradiography with (+)N-[11C]methyl-3-piperidyl benzilate in rats shows significant occupancy of brain muscarinic receptors with the intravenous injection of oxybutynin, solifenacin, and tolterodine. The estimated in vivo selectivity in brain is significantly greater for solifenacin and tolterodine than for oxybutynin. Imidafenacin occupies few brain muscarinic receptors. Similar findings for oral oxybutynin were observed with positron emission tomography in conscious rhesus monkeys with a significant disturbance of short-term memory. The newer generation of antimuscarinic agents may be advantageous in terms of bladder selectivity after systemic administration.
Administration, Cutaneous ; Animals ; Autoradiography ; Benzhydryl Compounds ; Brain ; Colon ; Cresols ; Heart ; Humans ; Imidazoles ; Injections, Intravenous ; Lung ; Macaca mulatta ; Mandelic Acids ; Memory, Short-Term ; Mice ; Muscarinic Antagonists ; Phenylpropanolamine ; Positron-Emission Tomography ; Quinuclidines ; Rats ; Receptors, Muscarinic ; Salivation ; Solifenacin Succinate ; Submandibular Gland ; Tetrahydroisoquinolines ; Tolterodine Tartrate ; Urinary Bladder ; Urinary Bladder, Overactive

PURPOSE: We investigated bladder function, with a special focus on nonvoiding contractions (NVCs), in awake rats with chronic chemical cystitis and bladder outlet obstruction (BOO) by use of simultaneous registrations of intravesical and intraabdominal pressures. In addition, we tested the effects of tolterodine on the NVCs in these models. METHODS: A total of 20 female Sprague-Dawley rats were used in this study. In eight rats, chemical cystitis was induced by intravesical instillation of HCl. Twelve rats were subjected to sham instillations or partial BOO. Four weeks after intravesical instillation or 2 weeks after partial BOO, cystometrograms were obtained by use of simultaneous recording of intravesical and intraabdominal pressure in all unanesthetized, unrestrained rats in metabolic cages. RESULTS: A total of 17 rats survived. In the rats with acute injury by HCl, 50% showed detrusor overactivity (DO), which was not seen in the sham group. The cystitis group had lower DO pressure without a difference in DO frequency compared with the BOO group. After the administration of tolterodine, the cystitis group showed no difference in DO frequency or pressure, whereas the BOO group showed decreased values for both parameters. CONCLUSIONS: Our study showed that toleterodine produced no effect on DO during the filling phase in rats with chronic chemical cystitisbut decreased the frequency and pressure of DO in rats with BOO. Clinically, studies are needed to improve the treatment effect of anticholinergic drugs ininterstitial cystitis patients with overactive bladder.
Administration, Intravesical ; Animals ; Benzhydryl Compounds ; Contracts ; Cresols ; Cystitis ; Female ; Humans ; Phenylpropanolamine ; Rats ; Rats, Sprague-Dawley ; Salicylamides ; Tolterodine Tartrate ; Urinary Bladder ; Urinary Bladder Neck Obstruction ; Urinary Bladder, Overactive ; Urodynamics

BACKGROUNDThe primary objectives of the treatment for the lower urinary tract symptoms (LUTS) associated with benign prostatic hyperplasia (BPH) are to produce rapid, sustained, and safe improvements in the symptoms that affect the quality of life in the majority of men over 50. In this study, we evaluated the efficacy and safety of the combined therapy with terazosin (apha1-adrenergic receptor antagonist) and tolterodine (anticholinergic agent) for LUTS associated with BPH.

METHODSThis combination study included 69 patients diagnosed with LUTS associated with BPH based on the International Prostate Symptom Scores (IPSS), urinary flow rate, prostate volume, urinary residual, and their serum prostate-specific antigen levels. Initially, 191 patients were treated with terazosin 2 mg once daily for one week. Those patients with continued LUTS after the initial treatment were allocated randomly into two groups: terazosin group (n = 36) in which patients were treated with terazosin 2 mg once daily for six weeks, and combination group (n = 33) in which patients were treated with both terazosin 2 mg once daily and tolterodine 2 mg twice daily for 6 weeks.

RESULTSThe IPSS were significantly improved in both groups after treatment, and the reduction of IPSS in the combination group was significantly greater than that in the terazosin group (P < 0.01). A decrease in urgency, frequency and nocturia were the main contributory factors causing the reduction of IPSS in the combination group. The differences about the peak urinary flow rate and the residual urine from the baseline values were noted in both groups after treatment, but were not significant between the two groups. The incidence of adverse effects in the combination group was higher than that in the terazosin group. As expected the most common adverse effect was mouth dryness which was associated with anticholinergic drugs such as tolterodine.

CONCLUSIONSPatients with LUTS associated BPH appear the improved IPSS after combined therapy with terazosin and tolterodine. This study, although short term and limited numbers of patients, provides evidence that the combined therapy with terazosin plus tolterodine is a good approach for meeting the objectives of rapid, sustained, and safe improvements in the LUTS associated with BPH. And the profile of patients in this study might be used as the indication of such combined therapy for LUTS associated with BPH without urodynamic evaluation.

Adrenergic alpha-Antagonists ; administration & dosage ; Aged ; Benzhydryl Compounds ; administration & dosage ; adverse effects ; Cresols ; administration & dosage ; adverse effects ; Drug Therapy, Combination ; Humans ; Male ; Middle Aged ; Muscarinic Antagonists ; administration & dosage ; Phenylpropanolamine ; administration & dosage ; adverse effects ; Prazosin ; administration & dosage ; adverse effects ; analogs & derivatives ; Prospective Studies ; Prostatic Hyperplasia ; complications ; drug therapy ; Tolterodine Tartrate ; Urination Disorders ; drug therapy