ObjectiveTo evaluate the feasibility of using near-infrared-Ⅱ (NIR-Ⅱ) rare-earth down-conversion nanoparticles (DCNPs) as a novel contrast agent for labeling adipose-derived mesenchymal stem cells (ADSCs), and to explore their potential application for in vivo tracking in a mouse model of subacute liver injury. MethodsCore-shell DCNPs (NaYF₄:Yb³⁺,Er³⁺@NaYF₄) were synthesized via a high-temperature thermal decomposition method. Water-soluble DCNPs were prepared through surface modification with DSPE-PEG2000, and further conjugated with fluorescein isothiocyanate (FITC) to observe cellular uptake. The CCK-8 assay was used to detect cell viability after co-incubation of ADSCs with DCNPs at 24 h and 48 h. Confocal laser scanning microscopy was employed to evaluate the labeling efficiency of DCNPs in ADSCs across various probe concentrations and incubation periods. Flow cytometry and differentiation assays were employed to evaluate the effects of DCNPs labeling on ADSCs surface antigen phenotype and multidirectional differentiation potential. NIR-Ⅱ fluorescence imaging was adopted to evaluate the dynamic changes of NIR‑Ⅱ fluorescence signals of labeled ADSCs during continuous in vitro culture. The in vivo toxicity of DCNP-labeled ADSCs transplanted via a single tail vein injection into SPF-grade male Sprague-Dawley (SD) rats was evaluated one month after transplantation through complete blood count, serum biochemical analysis, and multi-organ histopathological examination. A subacute liver injury model was induced in SPF-grade male BALB/c mice using carbon tetrachloride (CCl₄) administration. DCNP-labeled ADSCs were administered via tail vein injection, and the homing of ADSCs in vivo was monitored using NIR-Ⅱ fluorescence imaging. The effects of tracking on ADSC therapy were assessed using biochemical analysis of liver function indicators and hematoxylin and eosin (HE) staining. ResultsThe NaYF₄:Yb³⁺/Er³⁺ nanoparticles synthesized in this study exhibited a spherical crystal structure with particle sizes ranging from approximately 20 to 30 nm, and demonstrated good dispersion stability in water. CCK-8 results showed that DCNPs labeling exerted no cytotoxic effect on ADSCs viability after 24 and 48 h of labeling within the concentration range of 0 to 500 μg/mL used in this study. The cell labeling efficiency of DCNPs increased with dosage, while the cellular uptake remained essentially stable within 16 h. The NIR-Ⅱ fluorescence signal of DCNPs remained stable during the early stage of in vitro ADSCs culture. Although the fluorescence intensity decreased on day 3 of culture (P=0.003), detectable fluorescence signals were still observed in the cells on day 8. Flow cytometry and differentiation assays confirmed that DCNPs labeling had no significant adverse effects on the phenotype and differentiation capacity of ADSCs. One month after the transplantation of DCNP-labeled ADSCs into normal SD rats, blood routine tests, serum biochemical analysis, and multi-organ histopathological examination all revealed no obvious toxic reactions in the body. After the DCNP-labeled ADSCs were injected via the tail vein into mice with subacute liver injury, the serum levels of the liver function indicators alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were significantly reduced (both P<0.05), and the infiltration of inflammatory cells in the liver tissue was markedly decreased. Moreover, in vivo tracing results showed that the labeled ADSCs could home and migrate to the liver within 1 h, with the fluorescence signal gradually weakening over time. By 24 h, the fluorescence signal enriched in the liver had significantly decreased. ConclusionDCNPs possess good biocompatibility for ADSC labeling without affecting stem cell therapy. They may enable real-time, high signal-to-noise ratio in vivo tracking of stem cell homing and migration in ADSC therapy for subacute liver injury.

Objective To establish a rat model of type 2 diabetes mellitus with hepatic fibrosis using a high-lipid-sugar diet(HLSD)combined with inducement with carbon tetrachloride(CCl4)and streptozotocin.Methods SD rats were randomly divided into a liver fibrosis(LF)group,type 2 diabetes combined with liver fibrosis(DLF)group,and normal group,with 10 rats in each group.All rats in the LF or DLE group were fed an HLSD for 4 weeks;then,all rats were intraperitoneally injected with 20％CCl4(once a week)according to body weight(5 mL/kg)to induce hepatic fibrosis for another 4 weeks.During CCl4 treatment,rats in the DLF group were also intraperitoneally injected with 25 mg/kg STZ(once a week).After STZ treatment,the contents of FBG,INS,HOMA-IR,and HbA1c were evaluated once a week.The level of FBG was detected by blood glucose meter,and INS and HbA1c contents were detected by ELISA.Biochemical analysis was used to analyze the serum levels of AST,ALT,Tbil,and HA,as well as the HYP content in liver tissues.HE and Masson staining were used to evaluate pathological changes in liver tissues.Results The contents of FBG,HbA1c,and HOMA-IR were significantly increased,while the INS content was significantly decreased in the DLF group compared with those in the normal or LF group.Compared with the normal group,the LF group and DLF group showed higher levels of ALT,AST,Tbil,HA,and HYP,as well as fibrotic changes and inflammation in the liver tissue.Conclusions HLSD combined with CCl4 and STZ inducement is an effective strategy for establishing a rat model of type 2 diabetes mellitus with liver fibrosis.

Objective To establish a rat model of type 2 diabetes mellitus with hepatic fibrosis using a high-lipid-sugar diet(HLSD)combined with inducement with carbon tetrachloride(CCl4)and streptozotocin.Methods SD rats were randomly divided into a liver fibrosis(LF)group,type 2 diabetes combined with liver fibrosis(DLF)group,and normal group,with 10 rats in each group.All rats in the LF or DLE group were fed an HLSD for 4 weeks;then,all rats were intraperitoneally injected with 20％CCl4(once a week)according to body weight(5 mL/kg)to induce hepatic fibrosis for another 4 weeks.During CCl4 treatment,rats in the DLF group were also intraperitoneally injected with 25 mg/kg STZ(once a week).After STZ treatment,the contents of FBG,INS,HOMA-IR,and HbA1c were evaluated once a week.The level of FBG was detected by blood glucose meter,and INS and HbA1c contents were detected by ELISA.Biochemical analysis was used to analyze the serum levels of AST,ALT,Tbil,and HA,as well as the HYP content in liver tissues.HE and Masson staining were used to evaluate pathological changes in liver tissues.Results The contents of FBG,HbA1c,and HOMA-IR were significantly increased,while the INS content was significantly decreased in the DLF group compared with those in the normal or LF group.Compared with the normal group,the LF group and DLF group showed higher levels of ALT,AST,Tbil,HA,and HYP,as well as fibrotic changes and inflammation in the liver tissue.Conclusions HLSD combined with CCl4 and STZ inducement is an effective strategy for establishing a rat model of type 2 diabetes mellitus with liver fibrosis.

Objective To investigate the therapeutic effect of Liangxue-jiedu decoction on acute-on-chronic liver failure(ACLF)model mice and the Toll-like receptor 4(TLR4)/c-Jun amino terminal kinase(JNK)/nuclear factor κB(NF-κB)signaling pathway.Methods An ACLF mouse model was established using combined treatment of carbon tetrachloride,lipopolysaccharide,and D-galactosamine.Biochemical analysis was performed to evaluate liver function indicators,including alanine aminotransferase,aspartate aminotransferase,and total bilirubin.Histopathological examination was conducted to assess liver tissue morphological changes.Quantitative PCR was used to detect the mRNA expression of tumor necrosis factor alpha(TNF-α),interleukin(IL)-6,IL-1 β,and TLR4 in liver tissues.A CCK8 assay was used to evaluate the optimal interventional concentration of Liangxue-jiedu decoction on Raw 264.7 cells.Enzyme-linked immunosorbent assay was used to detect the contents of TNF-α,IL-6,and IL-1βin the cell supernatant.Protein immunoblotting was performed to measure the expression of TLR4/JNK/NF-κB signaling pathway-related proteins,including TLR4,NF-κB,p-ERKl/2,ERK1/2,p-JNK,and JNK.Results Compared with the ACLF model group,the Liangxue-jiedu decoction-treated group showed reduced cell necrosis,fibrosis,and inflammatory cell infiltration in liver tissues;decreased serum levels of alanine aminotransferase,aspartate aminotransferase and total bilirubin;and lower expression of TNF-α,IL-6,IL-1β,and TLR4 mRNA.Liangxue-jiedu decoction reduced TLR4/JNK/NF-κB signaling pathway-related protein expression in liver tissues.The in vitro result also showed that Liangxue-jiedu decoction reduced TNF-α,IL-6,and IL-1 β secretion by macrophage cells and down-regulated the expression of TLR4/JNK/NF-κB signaling pathway proteins.Conclusions Liangxue-jiedu decoction effectively improved liver function in ACLF mice in a manner closely related to the downregulation of TLR4/JNK/NF-κB pathway proteins.

BACKGROUND:Mineralized nodules are the mature marker of osteoblast differentiation, and the observation methods mainly use alizarin red staining.
OBJECTIVE:To compare the observation results of mineralized nodules by three methods, and to explore their characteristics and advantages, as wel as further application in the research of bone disease.
METHODS:The rat osteoblast-like cellline UMR-106 were cultured in the fresh medium that was changed every day, for 14 days. Alizarin red staining-light microscope, tetracycline fluorescence labeling-laser confocal scanning microscopy, and scanning electron microscopy were used to observe mineralized nodules. The calcium content of mineralized nodules was quantified using scanning electron microscopy and energy dispersive spectroscopy. In addition, tumor necrosis factor alpha that could inhibit the proliferation and differentiation of osteoblasts was used as the control.
RESULTS AND CONCLUSION:The three morphology methods could be used to observe the mineralized nodules of normal osteoblasts. As for tumor necrosis factor alpha, no mineralized nodules of osteoblasts were observed by alizarin red staining-light microscopy;smal mineralized nodules were observed by tetracycline staining-laser scanning confocal microscopy and scanning electron microscopy, suggesting tetracycline staining and scanning electron microscopy were more sensitive in the observation. Scanning electron microscopy could be used to observe the submicroscopic structures of mineralized nodules in the osteoblasts, and the formation of mineralized nodules, including the calcium secretion. Additional y, scanning electron microscopy combined with energy dispersive spectroscopy analysis can successful y quantify and position the mineralized nodules, indicating a potential application in the research of bone diseases.

BACKGROUND:Tougu Xiaotong capsule is the clinical prescription for the treatment of osteoarthritis in Fujian University of Traditional Chinese Medicine, and the previous studies mainly focus on effect to cartilage.
OBJECTIVE:To observe the effect of Tougu Xiaotong capsule on the proliferation and differentiation of osteoblasts as wel as the expressions of bone remodeling correlated factors.
METHODS:Rat osteoblast-like cel line ROS17/2.8 cel s were incubated with Tougu Xiaotong capsule. The ROS17/2.8 cel s were divided into blank control group and Tougu Xiaotong capsule groups with different
concentrations. The cel proliferation was determined by methylthiazolyldiphenyl-tetrazolium bromide assay. Osteoblast differentiation biomarkers alkaline phosphatase activity, osteocalcin and bone mineralized nodules were measured with colorimetry, enzyme-linked immunosorbent assay and alizarin red staining, respectively. The real-time PCR and enzyme-linked immunosorbent assay were used to detect the expressions of bone remodeling factors osteoprotegerin/receptor activator of nuclear factorκB ligand.
RESULTS AND CONCLUSION:Compared with the control group, the Tougu Xiaotong capsule with the
concentration of 0.25-2 g/L could significantly promote the ROS17/2.8 cel proliferation (P<0.05), up-regulate the alkaline phosphatase activity, osteocalcin expression level and mineralized nodules area, and increase the
percentage of bone remodeling factors osteoprotegerin/receptor activator of nuclear factorκB ligand (P<0.05). The mechanism of Tougu Xiaotong capsule protecting osteoarthritis may partly result from the regulation of
proliferation and differentiation of osteoblasts and bone remodeling.

Microscopic and histochemical methods were used to investigate flavonoids localization in the leaf and the stem of the Sarcandra glabra. The results indicated that flavonoids distributed mainly in epidermis, collenchyma, vascular bundles, secretory cells and palisade tissue of leaf. In the stem, they distributed mainly in epidermis, collenchyma, phloem and secretory cells. Histochemical localization of flavonoids using 5% solution of NaOH is convenient, rapid and reliable. The content of flavonoids in the leaf was higher those than in the stem. For sustainable utilization of the resources we suggested that only the leaves could be harvested.
Flavonoids ; metabolism ; Magnoliopsida ; metabolism ; Microscopy ; Plant Leaves ; metabolism ; Plant Stems ; metabolism