BACKGROUND:Previous studies have found that neuronal damage caused by continuous excessive fluoride exposure is related to Ca2+overload,but the mechanism of Ca2+flow conversion between intracellular calcium stores and cell apoptosis damage is still unclear.OBJECTIVE:To investigate the effect of fluoride exposure on Ca2+transport channel proteins and apoptosis levels in the mitochondria-associated endoplasmic reticulum membrane of primary cultured neural cells.METHODS:Primary nerve cells of neonatal SD rats were cultured in vitro and identified by immunofluorescence staining with neuronal nucleus-specific antibody up to day 7.The nerve cells were divided into control group(containing 0 mmol/L sodium fluoride),low fluoride group(containing 0.5 mmol/L sodium fluoride),and high fluoride group(containing 1 mmol/L sodium fluoride).The cell morphological changes were observed by light microscope 24 hours after fluorine exposure.The expression levels of apoptosis-related protein BAX/BCL-2 and calcium transfer-related pathways VDAC1,GRP 75,and IP3R were detected using western blot assay.The expression levels of VDAC1,GRP 75,and IP3R mRNA were detected by RT-PCR.Ca2+levels were detected by Rhood-2AM Ca2+probe.Mitochondrial membrane potential detection kit was used to detect the change in mitochondrial membrane potential.The level of apoptosis was determined by flow cytometry and TUNEL staining.RESULTS AND CONCLUSION:(1)The purity of neurons cultured on day 7 had been determined to be over 90％,with few impurities,good growth status,and tight cell network connections,meeting the requirements of subsequent experiments.(2)Compared with the control group,growth of neural cell clusters in the low-fluoride group and the high-fluoride group increased;the processes were broken;the cell body was rounded,and the connection network between cells was destroyed.Compared with the low-fluoride group,the cell damage changes in the high-fluoride group were more obvious.(3)Compared with the control group,the protein expressions of VDAC1,GRP75,and IP3R were increased in the low-fluoride group and the high-fluoride group(P＜0.05),and the ratio of apoptosis-related protein BAX/BCL-2 was increased(P＜0.05).Compared with the control group,the expression of VDAC1 and GRP75 mRNA in the low-fluoride group was significantly increased(P＜0.05);the expression levels of VDAC1,GRP75,and IP3R mRNA in the high-fluoride group were significantly increased(P＜0.01).(4)The level of cell apoptosis increased significantly after fluoride exposure,and the high-fluoride group was significantly higher than the control and low-fluoride groups(P＜0.01).(5)After fluoride exposure,the concentration of mitochondrial Ca2+in nerve cells increased significantly(P＜0.05),the mitochondrial membrane potential decreased(P＜0.01),and the degree of damage in the high-fluoride group was more obvious(P＜0.05).The results show that fluoride exposure impairs the morphological structure of primary neural cells,resulting in upregulation of Ca2+transfer pathway protein expression between the endoplasmic reticulum and mitochondria,mitochondrial Ca2+overload,mitochondrial damage,and increased levels of apoptosis.

BACKGROUND:Previous studies have found that neuronal damage caused by continuous excessive fluoride exposure is related to Ca2+overload,but the mechanism of Ca2+flow conversion between intracellular calcium stores and cell apoptosis damage is still unclear.OBJECTIVE:To investigate the effect of fluoride exposure on Ca2+transport channel proteins and apoptosis levels in the mitochondria-associated endoplasmic reticulum membrane of primary cultured neural cells.METHODS:Primary nerve cells of neonatal SD rats were cultured in vitro and identified by immunofluorescence staining with neuronal nucleus-specific antibody up to day 7.The nerve cells were divided into control group(containing 0 mmol/L sodium fluoride),low fluoride group(containing 0.5 mmol/L sodium fluoride),and high fluoride group(containing 1 mmol/L sodium fluoride).The cell morphological changes were observed by light microscope 24 hours after fluorine exposure.The expression levels of apoptosis-related protein BAX/BCL-2 and calcium transfer-related pathways VDAC1,GRP 75,and IP3R were detected using western blot assay.The expression levels of VDAC1,GRP 75,and IP3R mRNA were detected by RT-PCR.Ca2+levels were detected by Rhood-2AM Ca2+probe.Mitochondrial membrane potential detection kit was used to detect the change in mitochondrial membrane potential.The level of apoptosis was determined by flow cytometry and TUNEL staining.RESULTS AND CONCLUSION:(1)The purity of neurons cultured on day 7 had been determined to be over 90％,with few impurities,good growth status,and tight cell network connections,meeting the requirements of subsequent experiments.(2)Compared with the control group,growth of neural cell clusters in the low-fluoride group and the high-fluoride group increased;the processes were broken;the cell body was rounded,and the connection network between cells was destroyed.Compared with the low-fluoride group,the cell damage changes in the high-fluoride group were more obvious.(3)Compared with the control group,the protein expressions of VDAC1,GRP75,and IP3R were increased in the low-fluoride group and the high-fluoride group(P＜0.05),and the ratio of apoptosis-related protein BAX/BCL-2 was increased(P＜0.05).Compared with the control group,the expression of VDAC1 and GRP75 mRNA in the low-fluoride group was significantly increased(P＜0.05);the expression levels of VDAC1,GRP75,and IP3R mRNA in the high-fluoride group were significantly increased(P＜0.01).(4)The level of cell apoptosis increased significantly after fluoride exposure,and the high-fluoride group was significantly higher than the control and low-fluoride groups(P＜0.01).(5)After fluoride exposure,the concentration of mitochondrial Ca2+in nerve cells increased significantly(P＜0.05),the mitochondrial membrane potential decreased(P＜0.01),and the degree of damage in the high-fluoride group was more obvious(P＜0.05).The results show that fluoride exposure impairs the morphological structure of primary neural cells,resulting in upregulation of Ca2+transfer pathway protein expression between the endoplasmic reticulum and mitochondria,mitochondrial Ca2+overload,mitochondrial damage,and increased levels of apoptosis.

Patients with inflammatory bowel disease (IBD) require long-term pharmacotherapy. Due to the wide variety of clinically available drugs, medication safety has become increasingly concerning. As an indispensable component of pharmaceutical formulations, drug excipients may induce various adverse reactions, including rash, allergic responses, and gastrointestinal discomfort, and may even lead to severe organic injury. However, they are often overlooked in clinical practice. Therefore, identifying and managing excipient-related adverse reactions is crucial for ensuring medication safety. To address this issue, this article systematically reviews common excipients in IBD therapeutics, the underlying mechanisms of excipient-induced adverse reactions, and corresponding management strategies, aiming to enhance clinicians' awareness of excipient-related issues, improve their ability to recognize and manage such reactions, and ultimately ensure patient safety.

ObjectiveTo investigate the effect and mechanism of Shengxian Yixin Granule （升陷益心颗粒， SYG） in treating heart failure with preserved ejection fraction （HFpEF）. MethodsExperiment 1： Forty male C57BL/6 mice were randomly divided into control group， model group， low-dose ［25 mg/（kg·d）］， medium-dose ［50 mg/（kg·d）］， and high-dose ［（75 mg/（kg·d）］ SYG groups， with 8 mice per group. Except for the control group， HFpEF was induced by a high-fat diet combined with Nω-nitro-L-arginine methyl ester in drinking water for 16 weeks. After 10 weeks of modeling， treatment groups received corresponding doses of SYG by gavage， while the control group and model group received equal volume of normal saline， for 6 weeks. After 6 weeks of administration， the body weight， blood pressure including systolic blood pressure （SBP） and diastolic blood pressure （DBP）， and heart weight-to-tibia length ratio （HW/TL） were recorded. Echocardiography was used to evaluate left ventricular ejection fraction （LVEF） and the ratio of early to late diastolic transmitral flow velocity （E/A）. Plasma N-terminal pro-brain natriuretic peptide （NT-proBNP） and C-reactive protein （CRP） levels were measured by ELISA. The expression level of brain natriuretic peptide （BNP） in heart tissue was detected by qRT-PCR. Blood lipid levels， including low-density lipoprotein cholesterol （LDL-C）， triglycerides （TG）， and total cholesterol （TC）， were determined by a biochemical analyzer. Splenic proportions of M1 macrophages， M2 macrophages， T helper 17 （Th17） cells， and regulatory T （Treg） cells were analyzed by flow cytometry. Experiment 2： RAW 264.7 cells were stimulated with lipopolysaccharide （LPS） to establish an M1 macrophage polarization model， and then treated with SYG-containing serum for 24 h. The cells were divided into four groups， including blank serum group， high-dose SYG-containing serum group， LPS group， and LPS plus high-dose SYG-containing serum group. Combined with asignal transducer and activator of transcription 3 （Stat3） inhibitor for 24 h， the cells were divided into a lipopolysaccharide group， a high-dose SYG-containing serum group， an inhibitor group， and high-dose SYG-containing serum plus inhibitor group. The expression levels of inducible nitric oxide synthase （iNOS） and interleukin-1β （IL-1β） in each group were detected by qRT-PCR. The protein levels of iNOS， IL-1β， Stat3， and phosphorylated Stat3 （p-Stat3） were detected by Western Blotting. ResultsExperiment 1： Compared to the control group， the model group showed significantly decreased E/A ratio， and increased cardiac BNP expression， plasma NT-proBNP，HW/TL ratio， body weight， TC， LDL-C， SBP， and DBP， as well as shortened exhaustion distance and exhaustion time （P＜0.05 or P＜0.01）. Compared to the model group， the E/A ratio increased in the low-， medium-， and high-dose SYG groups； cardiac BNP expression decreased in the low- and high-dose groups， and plasma NT-proBNP levels decreased in the high-dose group； the HW/TL ratio and body weight decreased， while exhaustion distance and exhaustion time increased in the medium- and high-dose groups； TG levels decreased in the low- and medium-dose groups， and LDL-C levels decreased in the medium- and high-dose groups； SBP and DBP decreased in the low-， medium-， and high-dose groups （P＜0.05 or P＜0.01）. Compared to the low-dose SYG group， body weight decreased， and both exhaustion distance and exhaustion time increased in the medium- and high-dose groups （P＜0.05 or P＜0.01）. Compared to the medium-dose SYG group， the exhaustion time increased in the high-dose SYG group （P＜0.01）. Experiment 2： In the LPS‑induced M1 macrophage polarization model， compared to the blank serum group， the LPS group showed significantly increased mRNA and protein expression of iNOS and IL‑1β， as well as the p‑Stat3/Stat3 ratio （P＜0.01）. Compared to the LPS group， the high‑dose SYG-containing serum plus LPS group showed significantly decreased mRNA and protein expression of iNOS and IL‑1β， as well as the p‑Stat3/Stat3 ratio （P＜0.05 or P＜0.01）. Compared to the LPS group， the inhibitor group also showed significantly reduced these parameters （P＜0.05 or P＜0.01）. ConclusionSYG can improve cardiac function， ventricular remodeling， exercise capacity， and lipid metabolism in HFpEF mice， while inhibiting inflammatory responses. Its mechanism may be related to the regulation of the Stat3 signaling pathway and inhibition of M1 macrophage polarization.

ObjectiveTo investigate the effect and mechanism of Shengxian Yixin Granule （升陷益心颗粒， SYG） in treating heart failure with preserved ejection fraction （HFpEF）. MethodsExperiment 1： Forty male C57BL/6 mice were randomly divided into control group， model group， low-dose ［25 mg/（kg·d）］， medium-dose ［50 mg/（kg·d）］， and high-dose ［（75 mg/（kg·d）］ SYG groups， with 8 mice per group. Except for the control group， HFpEF was induced by a high-fat diet combined with Nω-nitro-L-arginine methyl ester in drinking water for 16 weeks. After 10 weeks of modeling， treatment groups received corresponding doses of SYG by gavage， while the control group and model group received equal volume of normal saline， for 6 weeks. After 6 weeks of administration， the body weight， blood pressure including systolic blood pressure （SBP） and diastolic blood pressure （DBP）， and heart weight-to-tibia length ratio （HW/TL） were recorded. Echocardiography was used to evaluate left ventricular ejection fraction （LVEF） and the ratio of early to late diastolic transmitral flow velocity （E/A）. Plasma N-terminal pro-brain natriuretic peptide （NT-proBNP） and C-reactive protein （CRP） levels were measured by ELISA. The expression level of brain natriuretic peptide （BNP） in heart tissue was detected by qRT-PCR. Blood lipid levels， including low-density lipoprotein cholesterol （LDL-C）， triglycerides （TG）， and total cholesterol （TC）， were determined by a biochemical analyzer. Splenic proportions of M1 macrophages， M2 macrophages， T helper 17 （Th17） cells， and regulatory T （Treg） cells were analyzed by flow cytometry. Experiment 2： RAW 264.7 cells were stimulated with lipopolysaccharide （LPS） to establish an M1 macrophage polarization model， and then treated with SYG-containing serum for 24 h. The cells were divided into four groups， including blank serum group， high-dose SYG-containing serum group， LPS group， and LPS plus high-dose SYG-containing serum group. Combined with asignal transducer and activator of transcription 3 （Stat3） inhibitor for 24 h， the cells were divided into a lipopolysaccharide group， a high-dose SYG-containing serum group， an inhibitor group， and high-dose SYG-containing serum plus inhibitor group. The expression levels of inducible nitric oxide synthase （iNOS） and interleukin-1β （IL-1β） in each group were detected by qRT-PCR. The protein levels of iNOS， IL-1β， Stat3， and phosphorylated Stat3 （p-Stat3） were detected by Western Blotting. ResultsExperiment 1： Compared to the control group， the model group showed significantly decreased E/A ratio， and increased cardiac BNP expression， plasma NT-proBNP，HW/TL ratio， body weight， TC， LDL-C， SBP， and DBP， as well as shortened exhaustion distance and exhaustion time （P＜0.05 or P＜0.01）. Compared to the model group， the E/A ratio increased in the low-， medium-， and high-dose SYG groups； cardiac BNP expression decreased in the low- and high-dose groups， and plasma NT-proBNP levels decreased in the high-dose group； the HW/TL ratio and body weight decreased， while exhaustion distance and exhaustion time increased in the medium- and high-dose groups； TG levels decreased in the low- and medium-dose groups， and LDL-C levels decreased in the medium- and high-dose groups； SBP and DBP decreased in the low-， medium-， and high-dose groups （P＜0.05 or P＜0.01）. Compared to the low-dose SYG group， body weight decreased， and both exhaustion distance and exhaustion time increased in the medium- and high-dose groups （P＜0.05 or P＜0.01）. Compared to the medium-dose SYG group， the exhaustion time increased in the high-dose SYG group （P＜0.01）. Experiment 2： In the LPS‑induced M1 macrophage polarization model， compared to the blank serum group， the LPS group showed significantly increased mRNA and protein expression of iNOS and IL‑1β， as well as the p‑Stat3/Stat3 ratio （P＜0.01）. Compared to the LPS group， the high‑dose SYG-containing serum plus LPS group showed significantly decreased mRNA and protein expression of iNOS and IL‑1β， as well as the p‑Stat3/Stat3 ratio （P＜0.05 or P＜0.01）. Compared to the LPS group， the inhibitor group also showed significantly reduced these parameters （P＜0.05 or P＜0.01）. ConclusionSYG can improve cardiac function， ventricular remodeling， exercise capacity， and lipid metabolism in HFpEF mice， while inhibiting inflammatory responses. Its mechanism may be related to the regulation of the Stat3 signaling pathway and inhibition of M1 macrophage polarization.

Intelligence encompasses various abilities, including logical reasoning, comprehension, self-awareness, learning, planning, creativity, and problem-solving. Extensive research and practical experience suggest that there are sex differences in intellectual development, with females typically maturing earlier than males. However, the key genes and molecular network mechanisms underlying these sex differences in intellectual development remain unclear. To date, Genome-Wide Association Studies (GWAS) have identified 507 genes that are significantly associated with intelligence. This study first analyzed RNA sequencing data from different stages of brain development (from BrainSpan), revealing that during the late embryonic stage, the average expression levels of intelligence-related genes are higher in males than in females, while the opposite is observed during puberty. This study further constructed interaction networks of intelligence-related genes with sex-differential expression in the brain, including the prenatal male network (HELP-M: intelligence genes with higher expression levels in prenatal males) and the pubertal female network (HELP-F: intelligence genes with higher expression levels in pubertal females). The findings indicate that the key genes in both networks are Ep300 and Ctnnb1. Specifically, Ep300 regulates the transcription of 53 genes in both HELP-M and HELP-F, while Ctnnb1 regulates the transcription of 45 genes. Ctnnb1 plays a more prominent role in HELP-M, while Ep300 is more crucial in HELP-F. Finally, this study conducted sequencing validation on rats at different developmental stages, and the results indicated that in the prefrontal cortex of female rats during adolescence, the expression levels of the intelligence genes in HELP-F, as well as key genes Ep300 and Ctnnb1, were higher than those in male rats. These genes were also involved in neurodevelopment-related biological processes. The findings reveal a sex-differentiated intelligence gene network and its key genes, which exhibit varying expression levels during the neurodevelopmental process.
Female ; Intelligence/physiology* ; Male ; Sex Characteristics ; Animals ; Brain/growth & development* ; E1A-Associated p300 Protein/physiology* ; beta Catenin/physiology* ; Transcriptome ; Rats ; Gene Expression Profiling ; Genome-Wide Association Study

Polygonati Rhizoma demonstrates significant potential for addressing both chronic and hidden hunger. The supply of high-quality seedlings is a primary factor influencing the development of the Polygonati Rhizoma industry. Warm water soaking is often used in agriculture to promote the rapid germination of seeds, while its application and molecular mechanism in Polygonati Rhizoma have not been reported. To rapidly obtain high-quality seedlings, this study treated Polygonatum kingianum var. grandifolium seeds with sand storage at low temperatures, warm water soaking, and cultivation temperature gradients. The results showed that the culture at 25 ℃ or sand storage at 4 ℃ for 2 months rapidly broke the seed dormancy of P. kingianum var. grandifolium, while the culture at 20 ℃ or sand storage at 4 ℃ for 1 month failed to break the seed dormancy. Soaking seeds in 60 ℃ warm water further increased the germination rate, germination potential, and germination index. Specifically, the seeds soaked at 60 ℃ and cultured at 25 ℃ without sand storage treatment(Aa25) achieved a germination rate of 78. 67%±1. 53% on day 42 and 83. 40%±4. 63% on day 77. The seeds pretreated with sand storage at 4 ℃ for 2 months, soaked in 60 ℃ water, and then cultured at 25 ℃ achieved a germination rate comparable to that of Aa25 on day 77. Transcriptomic analysis indicated that warm water soaking might promote germination by triggering reactive oxygen species( ROS), inducing the expression of heat shock factors( HSFs) and heat shock proteins( HSPs), which accelerated DNA replication, transcript maturation, translation, and processing, thereby facilitating the accumulation and turnover of genetic materials. According to the results of indoor controlled experiments and field practices, maintaining a germination and seedling cultivation environment at approximately 25 ℃ was crucial for the one-year seedling cultivation of P. kingianum var. grandifolium.
Germination ; Seedlings/genetics* ; Water/metabolism* ; Seeds/metabolism* ; Polygonatum/genetics* ; Temperature ; Plant Proteins/genetics* ; Plant Dormancy

This study aims to systematically characterize the targeted effects of Quanduzhong Capsules on cartilage lesions in knee osteoarthritis by integrating spatial transcriptomics data mining and animal experiments validation, thereby elucidating the related molecular mechanisms. A knee osteoarthritis model was established using Sprague-Dawley(SD) rats, via a modified Hulth method. Hematoxylin and eosin(HE) staining was employed to detect knee osteoarthritis-associated pathological changes in knee cartilage. Candidate targets of Quanduzhong Capsules were collected from the HIT 2.0 database, followed by bioinformatics analysis of spatial transcriptomics datasets(GSE254844) from cartilage tissues in clinical knee osteoarthritis patients to identify spatially specific disease genes. Furthermore, a "formula candidate targets-spatially specific genes in cartilage lesions" interaction network was constructed to explore the effects and major mechanisms of Quanduzhong Capsules in distinct cartilage regions. Experimental validation was conducted through immunohistochemistry using animal-derived biospecimens. The results indicated that Quanduzhong Capsules effectively inhibited the degenerative changes in the cartilage of affected joints in rats, which was associated with the regulation of Quanduzhong Capsules on the thioredoxin-interacting protein(TXNIP)-NOD-like receptor family pyrin domain containing 3(NLRP3)-bone morphogenetic protein receptor type 2(BMPR2)-fibronectin 1(FN1)-matrix metallopeptidase 2(MMP2) signal axis in the articular cartilage surface and superficial zones, subsequently inhibiting cartilage matrix degradation leading to oxidative stress and inflammatory diffusion. In summary, this study clarifies the spatially specific targeted effects and protective mechanisms of Quanduzhong Capsules within pathological cartilage regions in knee osteoarthritis, providing theoretical and experimental support for the clinical application of this drug in the targeted therapy on the inflamed cartilage.
Animals ; Osteoarthritis, Knee/metabolism* ; Drugs, Chinese Herbal/administration & dosage* ; Rats, Sprague-Dawley ; Rats ; Male ; Humans ; Capsules ; Female ; Disease Models, Animal