Serum Components Research Articles

Release of apoptosis inhibitor of macrophage (AIM) from pentameric IgM in serum predicts prognosis after hemodialysis initiation

BackgroundThe optimal timing for initiating dialysis and prognostic markers in chronic kidney disease (CKD) patients are under debate, with mortality and cardiovascular risks varying among patients. This study investigates whether the apoptosis inhibitor of macrophage (AIM), which is mostly bound to pentameric IgM, could serve as an effective indicator.MethodsWe prospectively followed 423 patients at dialysis initiation and 563 at various CKD stages. AIM dissociation from IgM and other serum components were measured in their serum samples. In vitro treatment of IgM-AIM complexes with their serum was conducted to assess AIM release from IgM. Survival analysis determined the associations of each variable with mortality and cardiovascular risk, and a cutoff value was calculated and validated using cross-validation.ResultsAIM dissociation from IgM increases with CKD progression and correlates with the serum uremic state, as shown by enhanced AIM release from IgM in vitro with sera from patients starting dialysis, but not those at earlier CKD stages. Patients at dialysis initiation with high proportion of serum IgM-free AIM (fAIM%) show elevated uremic toxins and other toxic metabolites, higher mortality, and increased cardiovascular risk compared to those with low fAIM%. This prognostic association is not seen with other CKD biomarkers, such as eGFR, creatinine, or inositol-phosphate. We determined the fAIM% cutoff of 46.27%, which predicts mortality two years post-dialysis initiation.ConclusionsThese findings suggest that the serum fAIM% could function as a prognostic marker at dialysis initiation and may have potential as a criterion for determining dialysis timing.

Optical sensing of albumin in human serum and urine-A historical review of the transition from classical dye-binding assays to advanced technologies.

Human serum albumin (HSA) is the most abundant protein in human plasma playing essential roles in transporting various biomolecules, metal ions, therapeutic agents, and metabolites. Additionally, it is crucial for maintaining oncotic pressure, scavenging free radicals, and preventing protein aggregation. Accurate quantification of HSA is vital for diagnosing various conditions, including hypertension, diabetes mellitus (DM), liver disorders, and renal diseases. While prevalent in clinical laboratories, traditional dye-binding methods have notable limitations: they can be time-consuming, lack sensitivity, and may suffer from interference from other serum components. These methods often require complex sample preparation and do not readily lend themselves to rapid or point-of-care testing (POCT). Consequently, there is a pressing need for innovative techniques that are rapid, cost-effective, and user-friendly. This review explores various dyes utilized for HSA determination, categorized into groups such as sulfonphthaleins, phenolphthaleins, azo dyes, etc., and provides a historical overview of the limitations of these methods. We critically assess the pros and cons of traditional dye-binding assays and emphasize the potential of emerging technologies, including microfluidic systems, smartphone-based detection, and nanopaper sensors, to address these gaps and enhance the efficiency and accessibility of HSA quantification in clinical settings.

The use of human albumin eye drops as an adjunctive therapy in cases of persistant corneal epithelial defects

Background Albumin eye drops (AED) can be effective in the healing of corneal epithelial defects. Albumin is the main protein component of autologous serum and therefore may be useful in the treatment of ocular surface diseases such as corneal ulcers and epithelial defects. Aim This work aimed to study the effect of human AED as adjunctive therapy in the treatment of persistent corneal epithelial defects (PCEDs). Patients and methods This prospective study was conducted in 20 patients aged 18–60 years, of both sexes, with PCEDs that did not respond to standard medical treatment for 10 to 14 days. All patients underwent slit-lamp examination, fundus examination, intraocular pressure measurement (digital), corneal swab, culture, and sensitivity. Results There was a significant positive correlation between the duration of medical therapy with the persistent antiepithelial defect AED and the size of the persistent epithelial defect at presentation (P<0.05). There was significant improvement in pain sensation, ciliary injection, persistent epithelial defect size, corneal, and stromal infiltration during the study period. There was no significant association between the duration of continuous ant epithelial drug therapy AED and stromal infiltration at presentation (P=0.627). Conclusion The promising therapeutic effects of AED on the corneal epithelium have been demonstrated. Albumin has shown a therapeutic effect in the treatment of PCEDs.

Modified Jiaoqi Powder enhances epithelial autophagy against TNF-triggered apoptosis in chronic ulcerative colitis

BackgroundA vicious cycle of dysregulated intestinal epithelial cell death, intestinal barrier defect, and subsequent inflammation response is core to chronic ulcerative colitis (UC). Modified Jiaoqi Powder (MJQP), a traditional Chinese medicine formula, has been clinically applied to treat chronic relapsing type and chronic persistent type of UC . Nevertheless, the underlying mechanism of MJPQ in chronic UC remains unknown. PurposeThe present study aimed to demonstrate the favorable effect and potential molecular mechanism of MJQP in chronic UC. MethodsThe chemical components of MJQP and MJQP drug serum were identified by LC-MS/MS. The curative effect of MJQP was evaluated in a well-established DSS-induced chronic UC mice model by measuring body weight, colon length, disease activity index (DAI) and histological scores . Serum cytokines, including interleukin (IL)-1β, IL-12, IL-13, IL-4, tumor necrosis factor-alpha (TNF-α) and INF-γ were measured using enzyme-linked immunosorbent assay. Western blotting, immunofluorescence, and MTT assay were used to analyze the effects of MJQP on colonic barrier function in chronic UC mice and human epithelial cell lines. TUNEL assay, western blotting, and flow cytometry were used to examine the related apoptosis indicators. An electron microscope was used to observe autophagosomes and autolysosomes, while western blotting and immunofluorescence were used to detect autophagy-associated proteins. Network pharmacology was used to predict potential targets and pathways of MJQP in UC. Finally, the TNF pathway-related proteins were detected by immunohistochemistry and western blotting. ResultsMJQP administration prevented the UC progression with faster weight gain, longer colon length, lower histological scores and DAI, and up/down- regulation of inflammatory factors. The expressions of tight junction proteins ki67 and E-cadherin increased dose-dependently after MJQP intervention. Moreover, MJQP treatment promoted the viability of NCM460 and Caco2 cells in a concentration-dependent manner. MJQP dose-dependently decreased the proportion of TUNEL-positive cells and attenuated the pro-apoptotic proteins cleaved-caspase 8 and cleaved-caspase 3 in colonic tissues. Flow cytometry also showed that MJQP dose-dependently decreased the apoptotic cell population of LPS-induced NCM460 and Caco2 cells. Electron microscopy revealed that autophagosomes and autolysosomes were significantly improved in the MJQP-treated groups. Additionally, autophagy-related proteins were significantly expressed after MJQP treatment. Network pharmacological analysis predicted that MJQP may alleviate chronic UC by affecting TNF-related signaling pathways and promoting intestinal epithelial cell proliferation. As anticipated, the TNF pathway-associated proteins were attenuated dose-dependently in colonic tissues after MJQP treatment. ConclusionThese results provide novel therapeutic strategies, indicating that MJQP may be a promising candidate treatment for chronic UC by promoting epithelial barrier restitution by enhancing epithelial autophagy against TNF–mediated apoptosis.

Lichong decoction improves inflammatory microenvironment and alleviates fibrosis in uterine leiomyoma via targeting CXCL8.

Lichong decoction (LD) is extensively employed in the treatment of uterine leiomyoma (ULM), demonstrating remarkable clinical effectiveness with an absence of notable adverse reactions. Its composition aligns with the traditional Chinese medicine (TCM) etiology of ULM, making it a highly suitable therapy. Nonetheless, the precise mechanisms underlying its therapeutic actions remain to be fully elucidated. The objective of this study was to clarify the therapeutic mechanism of LD improving ULM. The effects of LD on ULM cell viability, proliferation, and apoptosis were assessed using CCK-8, crystal violet staining, EdU incorporation, TUNEL, and Annexin V-FITC/PI assays. Gene microarray was used to profile differential gene expression after LD treatment. A rat ULM model was created to evaluate LD's anti-tumor efficacy, measuring body weight, uterine weight index, and sex hormone levels. Histopathological changes were analyzed with hematoxylin and eosin staining, Masson's trichrome staining, and transmission electron microscopy. Protein and RNA expression changes were analyzed via immunohistochemistry, western blotting, and qPCR. UHPLC-QE-MS enabled a detailed non-targeted LD analysis. Key components were identified through their correlation with serum sex hormones and inflammatory cytokines, and then examined by molecular docking studies. Experiments showed that LD reduced ULM cell viability and induced apoptosis. Gene expression profiling identified 313 differentially expressed genes. Enrichment analysis combined with experimental validation demonstrated that LD can reduce ULM fibrosis and inflammation by inhibiting the CXCL8/PI3K/AKT pathway. The analysis identified 494 primary compounds and 87 serum components in LD. Key compounds such as formononetin, palmatine, curcumenol, and hecogenin, which exhibit high affinity for CXCL8, may contribute to the anti-inflammatory and anti-tumor properties of LD. This study demonstrates that LD effectively inhibits ULM proliferation and fibrosis by improving the inflammatory microenvironment, primarily through the inhibition of CXCL8. These findings highlight the therapeutic potential of LD for ULM and provide new insights into its mechanisms.

A mammary gland secretion function exploratory study based on a quantitative comparison between milk and serum.

Lactation is a complex physiological process, and the knowledge about the secretion of many milk components is limited. The objective of this study was to explore the secretory function of human mammary glands through a comparative analysis of common clinical indicators in serum and milk. Milk and serum samples were collected from lactating women simultaneously, and titers of common biochemical components were determined using an automated biochemical analyzer. The differences between the levels of serum and milk components were analyzed using statistical software. There were significant biochemical differences between the milk and serum samples. Among the 46 biochemical components in the quantitative comparison, the titers or activities of nine milk constituents were significantly higher than those in serum, while 32 milk components were significantly lower than those in serum. Five constituents had significantly correlation between milk and serum. Overall, the individual differences in milk were significantly greater than those in serum. The differences between the levels of serum and milk components were assessed definitively, and some additional characteristic secretions from the human breast were also quantified, providing much more data for further research on the secretory function of the human breast.

Effect of water and feed quality on certain mineral elements (sodium, calcium, and chloride) in the blood serum of broiler males and females.

The study was carried out in one of the local fields in Nasiriyah District, Dhi Qar Governorate, from October 20 to November 25, 2023. To investigate the influence of feed and water quality on the concentration of certain mineral components in the blood serum of male and female broilers, I produced 300 one-day-old, sexed chicks and divided them into six treatments, each with three replicates. The study found that drinking water with RO had significantly higher levels of sodium (p≤0.05) than other water parameters. While there was no effect of the form of food or the sex of the broilers on the experimental parameters, Magnetic and RO water significantly outperformed Tap water in terms of calcium content in broiler blood serum (p≤0.05), Feed type and sex had no effect on the results of the study. The RO water treatment outperformed magnetized and Tap water treatments (p≤0.05) in terms of chloride content in male and female broiler blood serum. The form of feed and gender had no significant effect on the above-mentioned element concentration.

Assessing Cytotoxicity, Proteolytic Stability, and Selectivity of Antimicrobial Peptides: Implications for Orthopedic Applications.

In orthopedics, the use of anti-infective biomaterials is considered the most promising strategy to contrast the bacterial contamination of implant surfaces and reduce the infection rate. KSL, KSL-W, and Dadapin-1 are three antimicrobial peptides (AMPs) that possess significant antibacterial properties, making them promising candidates for producing anti-infective biomaterials not based on antibiotics. To fully assess their true potential, this study explores in detail their cytocompatibility on human osteoblast-like MG63 cells, murine fibroblastoid L929 cells, and hMSCs. To this end, the cytotoxicity of the AMPs in terms of IC50 was tested over a range of concentrations of 450-0.22 µg/mL using the ATP bioluminescence assay. The tests were performed both in the presence and absence of bovine serum to assess the effects of serum components on peptide stability. IC50 values obtained under the most stringent conditions were used to extrapolate the selectivity index (S.I.) toward salient bacterial species. In medium containing serum, all AMPs exhibited minimal to no cytotoxicity, with IC50 values exceeding 100 µg/mL. Dadapin-1 was the peptide that exhibited the lowest cytotoxicity, KSL-W exhibited the highest stability, and KSL exhibited the highest selectivity. Overall, these findings highlight the potential of these AMPs for the future production of anti-infective materials.

Chenodeoxycholic acid alleviated the cyclosporine-induced nephrotoxicity by decreasing oxidative stress and suppressing renin-angiotensin system through AT2R and ACE2 mRNA upregulation in rats.

Oxidative stress, inflammation and renin-angiotensin system (RAS) activation play an important role in the nephrotoxicity which is caused by the long-term use of the immunosuppressive drug cyclosporine (CsA). This study investigates whether chenodeoxycholic acid (CDCA), an endogenous farnesoid X receptor (FXR) agonist with antioxidant and anti-inflammatory effects, modulates CsA nephrotoxicity. CsA (25mg/kg/day; s.c.) was administered to rats for 12 days. CDCA (20mg/kg/day; i.p.) injection was started 3 days before CsA and continued for 15 days. CDCA improved renal damage and function in CsA-administered rats. Renal function markers in serum, renal histology, oxidative stress, inflammation and RAS components were determined in kidney. CDCA reduced CsA-induced renal increases in NADPH oxidase 4 and NADPH oxidase 2 mRNA expressions, oxidative stress and inflammation. CDCA elevated renal FXR, small heterodimer partner-1, hypoxia-inducible factor and vascular endothelial growth factor and nuclear factor erythroid 2-related factor mRNA expressions in CsA rats. It prevents renin angiotensin system activation by reducing angiotensin II (Ang-II) levels in serum and upregulating renal mRNA expressions of Ang II type-II receptor (AT2R) and angiotensin converting enzyme 2 (ACE2), but not AT1R and ACE in CsA rats. Our results indicate that CDCA may be a protective agent against CsA-nephrotoxicity by decreasing inflammation, oxidative stress and RAS activation via AT2R and ACE2 upregulations.

Effects of short-term salt exposure on gill damage, serum components and gene expression patterns in juvenile Largemouth bass (Micropterus salmoides)

The Largemouth bass (Micropterus salmoides; LMB) is a freshwater fish that plays a significant role in aquaculture, and its cultural base is expanding into inland saline water areas. To study the effect of short-term salt exposure on LMB, fish with an average body weight of 11.69 (±1.82) g were cultured for 14 days at three different salt concentrations (0 ‰, 6 ‰, and 12 ‰). After 14 days, the second gill arch was collected for tissue sectioning and transcriptome sequencing, while serum samples were collected to analyze serum components. The results showed that the mortality rate in the 0 ‰ and 6 ‰ groups was 0 %, whereas the mortality rate in the 12 ‰ group was 62 %. In the gill tissue sections, no apparent damage was observed in the 0 ‰ and 6 ‰ groups. However, in the 12 ‰ group, the secondary lamellae became shorter, thicker, and exhibited a disordered arrangement. The serum component test results showed that osmolality and K+ significantly increased in the 12 ‰ group, while Na+, K+, and Cl− concentrations showed slight increases, but the differences were not significant. Comparative transcriptome analysis revealed that, along the salinity gradient, gene expression exhibited five profiles. Genes related to ion transport and immunity were highly expressed in the 6 ‰ and 12 ‰ groups, while genes associated with biosynthesis and ATP production showed decreased expression levels as salinity increased. Notably, seven solute carrier genes, two Na+/K+-ATPase genes, and two insulin-like growth factor genes were significantly highly expressed in the 12 ‰ salinity group, playing important roles in the transmembrane transport of ions. Based on the results, the LMB can acclimatize to a salt concentration of at least 6 ‰. However, exposure to 12 ‰ salinity can lead to a series of adverse effects, including organ damage, reduced energy metabolism efficiency, and disruption of ion homeostasis.

Cremastrae Pseudobulbus Pleiones Pseudobulbus (CPPP) Against Non-Small-Cell Lung Cancer: Elucidating Effective Ingredients and Mechanism of Action.

Cremastrae Pseudobulbus Pleiones Pseudobulbus (CPPP) is derived from the dried pseudobulb of the orchid family plants Cremastra appendiculata (D.Don) Makino, Pleione bulbocodioides (Franch.) Rolfe, or Pleione yunnanensis Rolfe, and has the properties of clearing heat, detoxification, resolving phlegm, and dispersing nodules. It is frequently used for the treatment of various malignant tumors in clinical practice, especially lung cancer. CPPP is divided into two commercial specifications in the market, Maocigu (MCG) and Bingqiuzi (BQZ). However, owing to a lack of appropriate research strategies, the active ingredients and molecular mechanisms involved have not yet been clarified. This study intended to discover the combination of effective anti-lung-cancer ingredients in CPPP and explore their potential mechanisms of action. In this study, UHPLC-MS fingerprints of MCG and BQZ were established separately. Inhibitory effects on the proliferative viability and migratory ability of A459 and H1299 cells were evaluated as pharmacodynamic indicators. GRA and BCA were used to determine spectrum-effect relationships. Next, the identification and analysis of components of drug-containing serum were performed using UHPLC-Q-Exactive Orbitrap MS. Then, the results of the two analyses were combined to jointly screen out the anti-lung-cancer candidate active monomers of CPPP, and their in vitro activities were verified. Afterward, all effective ingredient combinations of MCG (MCGC) and BQZ (BQZC) were prepared according to their contents in the original medicinal materials. Their anti-lung-cancer activities in vitro and in vivo were compared and verified. Finally, we used the human lung cancer cell line A549 and the Lewis tumor xenograft model to investigate how BQZC would influence autophagy and apoptosis processes and the mechanisms involved. Overall, 11 predominant anti-lung-cancer active ingredients from CPPP were screened. Next, MCGC and BQZC were prepared according to their contents in the original medicinal materials, respectively, and their anti-tumor effects were equivalent to those of the original materials in vitro and in vivo. We found that BQZC could inhibit lung cancer cell growth and induce protective autophagy and apoptosis in lung cancer cells by activating the AMPK-mTOR-ULK1/BMF signaling pathway. These results provide important evidence for the clinical application and deep development of CPPP against tumors.

Quantitative analysis of dried serum FTIR spectra based on correlation Analysis-Interval random Frog-Partial least squares

Serum biochemical markers are widely used in clinical practice but often require expensive, specific reagents, complex instruments, and prolonged result waiting times. Infrared spectroscopy offers multiple advantages for serum analysis, such as reagent-free testing and the ability to quickly and directly measure multiple parameters simultaneously. This study collected serum samples from 66 healthy subjects to explore the relationship between dried serum infrared spectra and biochemical parameters, and to investigate the feasibility of simultaneously quantifying nine major serum components using dried serum infrared spectra. Initially, correlation analysis was conducted between spectral data and biochemical parameters, and the correlation spectral bands of glucose, protein and lipid were determined according to the correlation results. Subsequently, the interval random frog (IRF) algorithm was utilized to select the optimal characteristic wavenumbers of the correlated spectral bands, extracting the most informative spectral variables and constructing partial least squares (PLS) quantitative models. This method successfully achieved rapid and accurate quantification of nine major components in serum, including glucose, total protein, albumin, apolipoprotein A1, apolipoprotein B, total cholesterol, triglycerides, low-density lipoprotein cholesterol, and high-density lipoprotein cholesterol. The experimental results showed that the correlation coefficient (Rp) range in the test set was 0.8892–0.9941. Among them, the quantification of total cholesterol yielded the highest Rp, corresponding to a root mean square error (RMSEP) of 7.2425 mg/dL in the test set, while the quantification of glucose yielded the lowest Rp, with an associated RMSEP of 2.3683 mg/dL. The Correlation Analysis (CA)-IRF-PLS method developed in this study outperformed the conventional PLS method, the direct use of the successive projection algorithm (SPA)-PLS quantitative method and other reported quantitative techniques, providing a novel approach for the real-time determination of clinical parameters in serum.

Exploration the effective components of Gastrodia elata in improving cerebral ischemia reperfusion injury based on “Spectrum-effect” correlation and zebrafish verification experiment

BackgroundGastrodia elata (GE) has been widely used in clinical practice for many years with the functions of relieving stroke, suppressing liver Yang, dispelling wind and clearing collaterals. Our group's previous experimental studies have proved that GE has therapeutic effect on cerebral ischemia reperfusion injury (CIRI) (Ding et al., 2022). However, the active components of GE in treating CIRI remain unclear and require further research. PurposeThe purpose of this paper was to explore the potential effective components of GE improving CIRI based on the “Spectrum-effect” correlation. Zebrafish model was used for verification in vivo experimental. Materials and methodsFirst, the absorption components and metabolites of GE in rat serum were identified using ultra-high performance liquid chromatography-quadrupole time-of-flight tandem mass spectrometry (UHPLC-Q-TOF/MS). Second, pharmacodynamic indexes were determined by ELISA kit method, and the effect-time curve of each pharmacodynamic indexes was established. The potential compounds were screened using the statistical method of grey correlation between pharmacodynamic indicator and component response. Finally, the zebrafish CIRI model was successfully established, and the in vivo effect of the active components of GE was verified intuitively. Results45 chemical components were detected in GE. A total of 87 active components in serum of GE were identified including 25 prototype components and 62 metabolites. GE can improve CIRI by regulating the levels of interleukin-6 (IL-6), interleukin-1β (IL-1β), and tumor necrosis factor-α (TNF-α), MDA levels and SOD levels. It was found that p‑hydroxy benzaldehyde (PHB), p-hydroxybenzyl alcohol (PHBA) and gastrodin (GA) of GE were the possibly main active components by grey correlation statistics. The in vivo experiments of zebrafish model showed that PHB, PHBA, and GA have the ability to ameliorate cerebral thrombosis by regulation of oxidative stress and apoptosis. ConclusionsThe potential active components of GE on CIRI were initially excavated using UHPLC-Q-TOF-MS/MS, pharmacodynamics, and in vivo experiments of zebrafish model. It makes up for the disadvantages of separate research on chemical components and pharmacodynamics, and reflects the material basis of pharmacodynamics more objectively. It has provided theoretical basis for further quality evaluation and scientific foundation for rational drug using of GE in clinical.

Prevalence and Association of Components of Metabolic Syndrome and Uric Acid among Commercial Drivers in South-West Nigeria.

Metabolic syndrome (MetS) is a complex cluster of metabolic abnormalities characterized by central obesity, dyslipidemia, hypertension, and impaired glucose metabolism. Emerging evidence suggests a potential link between uric acid levels and MetS, and commercial drivers are exposed to unique occupational hazards that may predispose them to MetS and hyperuricemia. The objective of this study was to determine the prevalence of MetS and its relationship with serum uric acid among commercial drivers in Ado-Ekiti, Nigeria. A cross-sectional study was conducted among commercial drivers in Ado-Ekiti. Relevant information was obtained with a questionnaire, and anthropometry and blood pressure were measured. Fasting plasma glucose and lipid profiles were determined by the standard protocol. MetS was determined with the harmonized criteria. Bivariate correlation and Chi-square were used to determine the relationship between serum uric acid and MetS and its components. There were 106 participants with a median age (interquartile range) of 50 (11) years, with no difference between those with or without MetS. More than 70% of the participants drank alcohol and 75 (70.8%) participants had hyperuricemia. There was no significant correlation between serum uric acid and components of MetS. The prevalence of MetS among all the participants was 21.7% (23/106), with no difference among men with normal uric acid (25.8%) and those with elevated uric acid (20.0%), P = 0.509. Similarly, there was no statistical difference in the prevalence of the components of MetS between the two groups. The prevalence of MetS among the commercial drivers was high, with no difference among men with or without hyperuricemia. Given the importance of this group of people, a system-level public health approach should be adopted to promote a healthy lifestyle to save the lives of the populace.

Suppression of ferroptosis through the SLC7A11/glutathione/glutathione peroxidase 4 axis contributes to the therapeutic action of the Tangshenning formula on diabetic renal tubular injury

BackgroundTangshenning (TSN) is a safe and effective formula to treat diabetic nephropathy (DN), and clinical studies have demonstrated that its therapeutic effects are related to oxidative stress improvements in patients. Herein, this study aims to explore the potential mechanism of how TSN alleviates diabetic renal tubular injury.MethodsThe ultrahigh pressure liquid chromatography-quadrupole-time of flight mass spectrometry (UPLC-QTOF/MS) was used to identify the chemical composition and serum components of TSN. KK-Ay mice served to investigate the protective effects and regulatory mechanisms of TSN on tubular damage in DN. Furthermore, inhibitors and inducers of ferroptosis were employed in high glucose-cultured tubular epithelial cells (TECs) to verify the potential mechanisms of TSN. The expressions of proteins related to renal tubular injury, ferroptosis and solute carrier family 7, member 11 (SLC7A11)/glutathione (GSH)/glutathione peroxidase 4 (GPX4) axis were analyzed by western blot and immunofluorescence. Mitochondrial ultrastructure was observed in kidney tissues and TECs by a transmission electron microscope. Pathological changes in the renal tissues were observed by HE, PAS, and Prussian blue staining. Ferroptosis-related reactive oxygen species (ROS), malondialdehyde (MDA), ferrous ion, the intake of cystine, GSH, and oxidized glutathione (GSSG) were evaluated and contrasted in vivo or in vitro.Results51 compounds of TSN powder and 11 components in TSN-containing serum were identified by UPLC-QTOF/MS method. Administration of TSN ameliorated the elevated levels of proteinuria, serum creatinine, blood urea nitrogen, abnormal expression of renal tubular injury markers, and pathological damage to the renal tubules in DN mice model. Intriguingly, a strong inhibition of ferroptosis after TSN treatment occurred in both DN mice model and high glucose-cultured TECs. Notably, induction of ferroptosis by erastin attenuated the protective effect of TSN in high glucose-cultured TECs, while the ferroptosis inhibition by ferrostatin-1 treatment protected renal tubular, which was similar to TSN, suggesting the contribution of TSN-mediated by the inhibition of ferroptosis in DN progression. Mechanistically, TSN upregulated the SLC7A11/GSH/GPX4 axis to inhibit ferroptosis.ConclusionTSN may delay the DN progression and attenuate the renal tubular injury by inhibiting the ferroptosis regulated by the SLC7A11/GSH/GPX4 axis.

Huangqi-Danshen decoction improves heart failure by regulating pericardial adipose tissue derived extracellular vesicular miR-27a-3p to activate AMPKα2 mediated mitophagy

BackgroundHuangqi-Danshen decoction (HDD) is a classic traditional Chinese medicine for treating heart failure. Pericardial adipose tissue (PAT) has recently gained increasing attention in cardiovascular diseases. PurposeThis study aimed to investigate the effect of pericardial adipose tissue-derived extracellular vesicles on heart failure, the protective effect of HDD on myocardial remodel in heart failure rats, and identify the potential molecular mechanisms involved. MethodsUPLC-MS/MS identified active components of HDD. Extracellular vesicles (EVs) from pericardial adipose tissue of sham-operated and HF rats were identified through transmission electron microscopy, nanoparticle tracking analysis and western blot. EVs were co-cultured with H9c2 cardiomyocytes in order to examine their uptake and effects. MicroRNA sequencing, dual-luciferase reporter assay and PCR were conducted for exploring specific mechanisms of EVs on hypertrophic cardiomyocytes. In vivo, heart failure was modeled in rats via transverse aortic constriction (TAC). In vitro, the hypertrophic cardiomyocyte model were established using Ang II-induced H9c2 cardiomyocytes. ResultsUPLC-MS/MS identified 11 active components in serum of HDD administrated rats. Echocardiography showed HDD improved cardiac function in TAC model rats. HE and Masson staining indicated HDD ameliorated myocardial hypertrophy and fibrosis. MicroRNA sequencing found that HDD treatment resulted in 37 differentially expressed miRNAs (DMEs) (p < 0.05 and |log2FC| ≥ 1). KEGG analysis revealed that DEMs were enriched in the AMPK signaling pathway. PCR identified miR-27a-3p with the greatest difference in AMPK-related DMEs. Dual-luciferase reporter assay and Targetscan website were utilized to identify the target relationship between miR-27a-3p and PRKAA2 (AMPKα2). The miR-27a-3p negatively regulated AMPKα2 to inhibit mitophagy mediated by PINK1/Parkin pathway. HDD inhibited miR-27a-3p secretion from failing heart pericardial adipose tissue-derived extracellular vesicles, thereby improving inflammation, cardiac function, and myocardial remodeling through above pathways. ConclusionHDD inhibited the PAT-derived extracellular vesicular miR-27a-3p in failing hearts to activate AMPK/PINK1/Parkin signaling-mediated mitophagy, which improved cardiomyocyte energy metabolism, myocardial remodeling and heart failure.

Desmodium styracifolium (Osb.) Merr. Extracts alleviate cholestatic liver disease by FXR pathway

Ethnopharmacological relevanceCholestatic liver disease (CLD) is a disease characterized by cholestasis. Farnesoid X receptor (FXR) is a nuclear receptor that maintains homeostasis in bile acid metabolism. Studies have shown that gut microbiota interfered with the FXR pathway. Modulation of FXR to inhibit cholestasis has become a key measure in the treatment of CLD. In traditional folk medicine, Desmodium styracifolium (Osb.) Merr. was used as a primary treatment for gallstones, gonorrhea, jaundice, cholecystitis and other diseases. Modern pharmacological studies had also found that the herb has anti-calculus, anti-inflammatory, antioxidant, diuretic and liver damage. Therefore, we speculated that Desmodium styracifolium (Osb.) Merr. extracts (DME) could alleviate CLD through the FXR pathway and might be associated with the gut microbiota. However, studies of DME alleviating CLD through the FXR pathway have not been reported. Aim of studyTo study the effect and mechanism of DME in relieving CLD through in vivo and in vitro experiments. Materials and methodsFirst, mice were administrated with alpha-naphthyl isothiocyanate (ANIT) to establish a CLD model in vivo. Meanwhile, HepG2 cells were induced by lithocholic acid (LCA) to establish the CLD model in vitro. To evaluate the therapeutic effect of DME on CLD mice, hematoxylin-eosin (HE) staining, and biochemical indicators were performed. The prototype of the blood components in mice serum was detected by ultra-high-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). 16S rDNA sequencing was used to analyze the gut microbiota. Finally, the protein and mRNA expression of the FXR pathway in mice liver tissues or HepG2 cells were detected by Western blot, qRT-PCR, or immunofluorescence. ResultsPathological testing and biochemical indexes showed that DME significantly reduced serum ALT, AST, ALP, TBIL, DBIL, TBA and liver TBA levels, and attenuated liver tissue injury, necrosis and jaundice in CLD mice. In addition, MetagenomeSeq analysis of gut microbiota showed that DME significantly up-regulated the abundance of Parvibacter, down-regulated the abundance of Paenalcaligenes, and regulated bile acid homeostasis. In terms of mRNA expression, DME significantly upregulated the mRNA levels of Nr1h4, Abcb11, Cyp7a1 and Slc10a1. Meanwhile, in terms of protein expression, DME significantly up-regulated the protein expression levels of FXR, BSEP, CYP7A1 and NTCP, which regulated bile acid homeostasis. Finally, the molecular docking results showed that the components of DME, such as Lumichrome, Daidzein and Folic acid, all had good binding ability with FXR, and the surface plasmon resonance (SPR) results also showed that both Lumichrome and Daidzein had a relatively high affinity with FXR. ConclusionDME alleviated CLD through the FXR pathway, and the mechanisms might be associated with the gut microbiota.

Therapeutic effects and mechanisms of Modified Ma-Xing-Shi-Gan Decoction on Klebsiella pneumoniae-induced pneumonia in mice assessed by Multi-omics

Ethnopharmacological relevanceModified Ma-Xing-Shi-Gan decoction (MMXSGD), a classic prescription from Treatise on Febrile Disease in China, is commonly used to treat Klebsiella pneumoniae (KP) infections in clinical settings. Materials and methodsThe aim of this study is to assess the efficacy of MMXSGD in the treatment of pneumonia and investigate its underlying mechanism of action. UHPLC-MS/MS was established to identify the main chemical components of serum after intragastric administration with MMXSGD. A mouse model of pneumonia caused by KP was used to evaluate the therapeutic potential of MMXSGD. The macrophage polarization was analyzed by immunohistochemistry. The cytokine profile was assessed using Luminex assay. Lung transcript and metabolite levels were assessed by transcriptomics and non-targeted metabolomics to analyze potential anti-pneumonia mechanisms and targets. Results22 major blood-entry components and 274 MMXSGD-pneumonia-related targets were identified. Compared with the model group, the mortality rate of mice in different dosage groups of MMXSGD was significantly reduced, and pathological lung damage was significantly alleviated. Among them, the low dose of MMXSGD treatment had the best protective effect. Further, MMXSGD treatment could regulates M1/M2 polarization in macrophages and inhibits the production of pro-inflammatory cytokines. The data from transcriptome and metabolome analysis indicate that MMXSGD could regulate inflammation-related pathways (PI3K/AKT, HIF-1, NF-κB pathway) and metabolites to modulate pulmonary inflammation. The results demonstrate that MMXSGD enhances the antibacterial effect in vivo by suppressing inflammation and regulating immunity rather than directly antibacterial effect. ConclusionThese findings provide a further assessment of MMXSGD, suggesting that MMXSGD has good therapeutic efficacy in bacterial infectious diseases.

Tongsai Granules inhibit autophagy and macrophage-mediated inflammatory response to improve acute exacerbations of chronic obstructive pulmonary disease in rats

To investigate the inhibitory effect of Tongsai Granules (TSG) on macrophage-mediated inflammatory response to alleviate acute exacerbation of chronic obstructive pulmonary disease (AECOPD) in rats and explore the underlying mechanism. Twenty-four rats were divided into control group, AECOPD model group, TSG treatment group, and moxifloxacin+salbutamol (MXF+STL) treatment group. In the rat models of COPD, AECOPD was induced by nasal instillation of Klebsiella pneumoniae on day 3 of week 9 after modeling, and saline, TSG or MXF+STL were administered via gavage on days 1 and 2 and days 4 to 7 of week 9. After the treatments, lung tissues were collected for examining for pathologies and expressions of inflammatory markers, MMP2, and MMP9. In cultured macrophage MH-S cells with LPS stimulation, the effect of TSG-medicated serum on IL-1β, IL-6, TNF-α, COX-2, and iNOS expressions and phosphorylation levels of p38, p-p62, LC3, FoxO3a, and mTOR were evaluated. TSG significantly improved lung pathologies and lung function in AECOPD rats by reducing bronchial wall thickness and mean alveolar linear intercept, increasing alveolar numbers, and reducing pulmonary expression of IL-1β, IL-6, TNF- α, MMP2 and MMP9. In MH-S cells, TSG significantly suppressed LPS-induced expressions of inflammatory cytokines, COX-2 and iNOS. Serum pharmacology coupled with network pharmacology identified 10 chemical components in TSG-medicated serum, and functional analysis of their 466 targets suggested that the therapeutic effect of TSG on AECOPD was mediated primarily by luteolin and quercetin, which regulate the MAPK, mTOR, FoxO, and autophagy pathways. In MH-S cells, luteolin significantly inhibited LPS-induced inflammatory responses and expressions of p-p38, FoxO3a, mTOR, p-p62 and LC3. TSG reduces macrophage-mediated inflammatory responses to alleviate AECOPD in rats possibly by modulating p38, mTOR, and FoxO3a pathways and inhibiting autophagy.

Effective Components and Molecular Mechanism of Biejiaruangan Capsule Against Liver Fibrosis: High-resolution Mass Spectrometry, Network Pharmacological Analysis and Experimental Verification.

Biejiaruangan capsule (BJRGC) is a commonly used traditional Chinese medicine preparation for treating oftreating liver fibrosis (LF), but its specific molecular mechanism is unclear. This study used mass spectrometry, network pharmacology and experimental verification to explore the mechanism of BJRGC against LF. Ultrahigh-performance liquid chromatography-quadrupole-exactive-orbitrap-mass spectrometry (UHPLC-Q-Exactive-Orbitrap-MS) and network pharmacology were employed to identify and screen the potential components, targets, and signaling pathways of BJRGC against LF. The interaction between the active ingredients and targets was validated using molecular docking. Finally, 5-ethynyl-2'-deoxyuridine (EDU) staining, western blotting (WB), and flow cytometry (FCM) were utilized to further verify the mechanism of BJRGC against LF. A total of 9 prototype components of BJRGC were identified in serum, most derived from iridoid glycosides and triterpenes in Gardenia jasminoides Ellis and Artemisia scoparia Waldst.et Kit. Network pharmacology predicts that medicine prototype components in serum mostly influence targets such as CDK2, CDK6, and PIK3CG, with the key route being the PI3K/AKT signaling pathway. Molecular docking showed that the major components have good binding properties with key target proteins. The experimental results showed that BJRGC could inhibit the proliferation of HSCs, induce cell cycle arrest and reduce the protein expression of CDK2, CDK6 and PIK3CG. BJRGC can inhibit the proliferation of HSCs by targeting the protein expression of CDK2, CDK6, and PIK3CG in the PI3K/AKT signaling pathway through its prototype components, such as hyperoside, tumulosic acid, and hederagenin, thereby alleviating LF disease.