This paper aimed to explore the effects of Duzhong Decoction(DZD)-containing serum on the proliferation and osteoblast differentiation of MC3T3-E1 cells through L-type voltage-gated calcium channels(L-VGCCs). L-VGCCs inhibitors, nifedipine and verapamil, were used to block L-VGCCs in osteoblasts. MC3T3-E1 cells were divided into a control group, a low-dose DZD-containing serum(L-DZD) group, a medium-dose DZD-containing serum(M-DZD) group, a high-dose DZD-containing serum(H-DZD) group, a nifedipine group, a H-DZD + nifedipine group, verapamil group, and a H-DZD + verapamil group. The CCK-8 method was used for cell proliferation analysis, alkaline phosphatase(ALP) assay kits for intracellular ALP activity measurement, Western blot for protein expression level in cells, real-time fluorescence quantitative PCR technology for intracellular mRNA expression level determination, fluorescence spectrophotometer for free Ca~(2+) concentration determination in osteoblasts, and alizarin red staining(ARS) for mineralized nodule formation in osteoblasts. The experimental results show that compared to the control group, DZD groups can promote MC3T3-E1 cell proliferation, ALP activity, and mineralized nodule formation, increase intracellular Ca~(2+) concentrations, and upregulate the protein expression of bone morphogenetic protein 2(BMP2), collagen Ⅰ(COL1), α2 subunit protein of L-VGCCs(L-VGCCα2), and the mRNA expression of Runt-related transcription factor 2(RUNX2), and BMP2. After blocking L-VGCCs with nifedipine and verapamil, the intervention effects of DZD-containing serum were inhibited to varying degrees. Both nifedipine and verapamil could inhibit ALP activity, reduce mineralized nodule areas, and downregulate the expression of bone formation-related proteins. Moreover, the effects of DZD-containing serum on increasing MC3T3-E1 cell proliferation, osteoblast differentiation, and Ca~(2+) concentrations, upregulating the mRNA expression of osteoprotegerin(OPG) and protein expression of phosphorylated protein kinase B(p-Akt) and phosphorylated forkhead box protein O1(p-FOXO1), and upregulating phosphatase and tensin homolog(PTEN) expression were reversed by nifedipine. The results indicate that DZD-containing serum can increase the Ca~(2+) concentration in MC3T3-E1 cells to promote bone formation, which may be mediated by L-VGCCs and the PTEN/Akt/FoxO1 signaling pathway, providing a new perspective on the mechanism of DZD in treating osteoporosis.
Animals ; Osteoblasts/metabolism* ; Cell Proliferation/drug effects* ; Cell Differentiation/drug effects* ; Mice ; Drugs, Chinese Herbal/pharmacology* ; Calcium Channels, L-Type/genetics* ; Alkaline Phosphatase/genetics* ; Serum/chemistry* ; Cell Line ; Osteogenesis/drug effects* ; Bone Morphogenetic Protein 2/genetics*

OBJECTIVE: To explore the anti-photoaging properties of salvianolic acid B (Sal B). METHODS: The optimal photoaging model of human immortalized keratinocytes (HaCaT cells) were constructed by expose to ultraviolet B (UVB) radiation. The cells were divided into control, model and different concentrations of Sal B groups. Cell viability was measured via cell counting kit-8. Subsequently, the levels of oxidative stress, including reactive oxygen species (ROS), hydroxyproline (Hyp), catalase (CAT), and glutathione peroxidase (GSH-Px) were detected using the relevant kits. Silent information regulator 1 (SIRT1) protein level was detected using Western blot. The binding pattern of Sal B and SIRT1 was determined via molecular docking. RESULTS: Sal B significantly increased the viability of UVB-irradiated HaCaT cells (P<0.05 or P<0.01). Sal B effectively scavenged the accumulation of ROS induced by UVB (P<0.05 or P<0.01). In addition, Sal B modulated oxidative stress by increasing the intracellular concentrations of Hyp and CAT and the activity of GSH-Px (P<0.05 or P<0.01). The Western blot results revealed a substantial increase in SIRT1 protein levels following Sal B administration (P<0.05). Moreover, Sal B exhibited good binding affinity toward SIRT1, with a docking energy of -7.5 kCal/mol. CONCLUSION Sal B could improve the repair of photodamaged cells by alleviating cellular oxidative stress and regulating the expression of SIRT1 protein.
Humans ; Sirtuin 1/metabolism* ; Ultraviolet Rays ; Oxidative Stress/radiation effects* ; Keratinocytes/metabolism* ; Molecular Docking Simulation ; Benzofurans/pharmacology* ; Skin Aging/radiation effects* ; Reactive Oxygen Species/metabolism* ; Cell Survival/radiation effects* ; HaCaT Cells ; Hydroxyproline/metabolism* ; Glutathione Peroxidase/metabolism* ; Catalase/metabolism* ; Depsides

Patients suffering from nerve injury often experience exacerbated pain responses and complain of memory deficits. The dorsal hippocampus (dHPC), a well-defined region responsible for learning and memory, displays maladaptive plasticity upon injury, which is assumed to underlie pain hypersensitivity and cognitive deficits. However, much attention has thus far been paid to intracellular mechanisms of plasticity rather than extracellular alterations that might trigger and facilitate intracellular changes. Emerging evidence has shown that nerve injury alters the microarchitecture of the extracellular matrix (ECM) and decreases ECM rigidity in the dHPC. Despite this, it remains elusive which element of the ECM in the dHPC is affected and how it contributes to neuropathic pain and comorbid cognitive deficits. Laminin, a key element of the ECM, consists of α-, β-, and γ-chains and has been implicated in several pathophysiological processes. Here, we showed that peripheral nerve injury downregulates laminin β1 (LAMB1) in the dHPC. Silencing of hippocampal LAMB1 exacerbates pain sensitivity and induces cognitive dysfunction. Further mechanistic analysis revealed that loss of hippocampal LAMB1 causes dysregulated Src/NR2A signaling cascades via interaction with integrin β1, leading to decreased Ca²⁺ levels in pyramidal neurons, which in turn orchestrates structural and functional plasticity and eventually results in exaggerated pain responses and cognitive deficits. In this study, we shed new light on the functional capability of hippocampal ECM LAMB1 in the modulation of neuropathic pain and comorbid cognitive deficits, and reveal a mechanism that conveys extracellular alterations to intracellular plasticity. Moreover, we identified hippocampal LAMB1/integrin β1 signaling as a potential therapeutic target for the treatment of neuropathic pain and related memory loss.
Animals ; Laminin/genetics* ; Hippocampus/metabolism* ; Neuralgia/metabolism* ; Cognitive Dysfunction/etiology* ; Male ; Peripheral Nerve Injuries/metabolism* ; Extracellular Matrix/metabolism* ; Integrin beta1/metabolism* ; Pyramidal Cells/metabolism* ; Signal Transduction

Sennoside A (SA), a typical prodrug, exerts its laxative effect only after its transformation into rheinanthrone catalyzed by gut microbial hydrolases and reductases. Hydrolases have been identified, but reductases remain unknown. By linking a photoreactive group to the SA scaffold, we synthesized a photoaffinity probe to covalently label SA reductases and identified SA reductases using activity-based protein profiling (ABPP). From lysates of an active strain, Bifidobacterium pseudocatenulatum (B. pseudocatenulatum), 397 proteins were enriched and subsequently identified using mass spectrometry (MS). Among these proteins, chromate reductase/nicotinamide adenine dinucleotide (NADH) phosphate (NADPH)-dependent flavin mononucleotide (FMN) reductase/oxygen-insensitive NADPH nitroreductase (nfrA) was identified as a potent SA reductase through further bioinformatic analysis and The Universal Protein Resource (UniProt) database screening. We also determined that recombinant nfrA could reduce SA. Our study contributes to further illuminating mechanisms of SA transformation to rheinanthrone and simultaneously offers an effective method to identify gut bacterial reductases.

Mercury (Hg)-contaminated soil poses a significant threat to the environment and human health. Hg-resistant microorganisms have the ability to survive under the stress of inorganic and organic Hg and effectively reduce Hg levels and toxicity. Compared to physical and chemical remediation methods, microbial remediation technologies have garnered increasing attention in recent years due to their lower cost, remarkable efficacy, and minimal environmental impact. This paper systematically elucidates the molecular mechanisms of Hg resistance in microbes, with a focus on their potential applications in phytoremediation of Hg-contaminated soils through plant-microbe interactions. Furthermore, it highlights the critical role of microbes in enhancing the effectiveness of transgenic plants for Hg remediation, aiming to provide a theoretical foundation and scientific basis for the bioremediation of Hg-contaminated soils.
Mercury/toxicity* ; Biodegradation, Environmental ; Soil Pollutants/isolation & purification* ; Soil Microbiology ; Plants, Genetically Modified/metabolism* ; Bacteria/genetics*

OBJECTIVE: To study the in vitro and in vivo antitumor effects of the polysaccharide of Alocasia cucullata (PAC) and the underlying mechanism. METHODS: B16F10 and 4T1 cells were cultured with PAC of 40 µg/mL, and PAC was withdrawn after 40 days of administration. The cell viability was detected by cell counting kit-8. The expression of Bcl-2 and Caspase-3 proteins were detected by Western blot and the expressions of ERK1/2 mRNA were detected by quantitative real-time polymerase chain reaction (qRT-PCR). A mouse melanoma model was established to study the effect of PAC during long-time administration. Mice were divided into 3 treatment groups: control group treated with saline water, positive control group (LNT group) treated with lentinan at 100 mg/(kg·d), and PAC group treated with PAC at 120 mg/(kg·d). The pathological changes of tumor tissues were observed by hematoxylin-eosin staining. The apoptosis of tumor tissues was detected by TUNEL staining. Bcl-2 and Caspase-3 protein expressions were detected by immunohistochemistry, and the expressions of ERK1/2, JNK1 and p38 mRNA were detected by qRT-PCR. RESULTS: In vitro, no strong inhibitory effects of PAC were found in various tumor cells after 48 or 72 h of administration. Interestingly however, after 40 days of cultivation under PAC, an inhibitory effect on B16F10 cells was found. Correspondingly, the long-time administration of PAC led to downregulation of Bcl-2 protein (P<0.05), up-regulation of Caspase-3 protein (P<0.05) and ERK1 mRNA (P<0.05) in B16F10 cells. The above results were verified by in vivo experiments. In addition, viability of B16F10 cells under long-time administration culture in vitro decreased after drug withdrawal, and similar results were also observed in 4T1 cells. CONCLUSIONS Long-time administration of PAC can significantly inhibit viability and promote apoptosis of tumor cells, and had obvious antitumor effect in tumor-bearing mice.
Mice ; Animals ; Alocasia/metabolism* ; MAP Kinase Signaling System ; Caspase 3/metabolism* ; Apoptosis ; RNA, Messenger/metabolism*

Perioperative neurocognitive disorder(PND)significantly threatens brain health,leading to prolonged hospitalization,increased patient mortality risk,and poor long-term prognosis.Sleep disorder may substantially elevate the risk of neurocognitive dysfunction.However,the specific role of sleep in PND development remains to be elucidated.In addition,controversy exists over whether interventions for perioperative sleep disorder can effectively prevent PND.Therefore,this review aims to explore the basic function and regulatory principles of sleep,the clinical characteristics of perioperative sleep disorder,its role and potential mechanisms in PND development.Furthermore,we also provide new evidence on the potential therapeutic strategies to modulate sleep disorder for PND management,to improve patient's neurocognitive prognosis and long-term outcomes.

Objective:To investigate the association of urinary cadmium levels with peripheral leukocyte classification counts among middle-aged and older adults aged 40 to 89 years in selected areas of China.Methods:The research was based on the survey of the impact of soil quality of agricultural land on human health in typical areas conducted in 2019-2020. A total of 5 600 middle-aged and older adults aged 40 to 89 years were included by using a multi-stage stratified random sampling method. Baseline characteristics of the subjects were collected and physical examinations were performed. Random midstream urine was collected to measure urinary cadmium and urinary creatinine and fasting venous blood was collected to measure the leukocyte count, neutrophil count, lymphocyte count, monocyte count and eosinophil count. The linear mixed effect model was used to analyse the association of urinary cadmium levels with leukocyte classification counts, and the dose-response relationship between them was analyzed by using the restricted cubic spline (RCS) function.Results:The age of the subjects was (63.17±12.02) years; 2 851 (50.91%) were males; and the M ( Q 1, Q 3) of urinary creatinine-corrected urinary cadmium levels was 2.69 (1.52, 4.69) μg/g·creatinine. After adjusting for confounding factors, the results of linear mixed effects model analysis showed that for each 1-unit increase in urinary creatinine-corrected urinary cadmium level, the percentage change [% (95% CI)] of leukocyte count and lymphocyte count was -1.70% (-2.61%, -0.79%) and -1.57% (-2.86%, -0.26%), respectively. RCS function showed a negative linear relationship between urinary creatinine-corrected urinary cadmium levels and leukocyte counts and lymphocyte counts, respectively (all Pnon-linear>0.05). Conclusion:Urinary cadmium levels are negatively associated with leukocyte count and lymphocyte count among middle-aged and older adults aged 40 to 89 years in selected areas of China.

Objective To determine whether four-and-a-half LIM-only protein 2(FHL2)can affect macrophage foa-ming by regulating nuclear factor kappa-B(NF-κB)signaling pathway.Methods FHL2 over-expression plasmids and siRNA of FHL2 were constructed and transfected into human monocyte/macrophages cell line THP-1.Western blot was used to detect the expression of FHL2.The cells were stimulated with oxidized low density lipoprotein(ox-LDL)and the expression of IL-6,IL-1β,TNF-α and other cytokines were detected by ELISA.Oil red O staining was used to detect the degree of cell foaming.The protein expression of NF-κB signaling pathway was detected by Western blot.Results The expression of FHL2 increased after transfected with FHL2 over-expression plasmids while decreased in si-FLH2 transfected cells.FHL2 down-regulated secretion of inflammatory cytokines.Down-regulation of FHL2 allevi-ated THP-1 macrophage foaming.The down-regulation of FHL2 inhibited activation of NF-κB signaling pathway,while the over-expression FHL2 showed an opposite trend.Conclusions FHL2 down-regulation inhibits the activation of NF-κB signaling pathway,reduces the secretion of inflammatory cytokines and alleviates foaming of macrophages.

With the increasing proportion of laboratory projects in the revenue of public hospitals,how to adjust the prices of labo-ratory projects and improve the income structure of public hospitals has become one of the important contents of China's medical re-form.It is based on a systematic introduction and summary of the pricing methods for inspection projects in British Columbia,Canada,the US,and WHO.Combining with China's national conditions,it proposes three policy recommendations:establishing a cost cal-culation system and cost calculation tools for inspection projects,adopting differentiated pricing strategies based on social medical secu-rity levels,and regularly reviewing and updating the prices of inspection projects.