Objective:To investigate the effect of β-Ecdysterone (β-Ecd), an active ingredient in cow′s knee, on hormonal femoral head necrosis in young rabbits and explore the mechanism involved.Methods:An animal study.Sixty New Zealand young rabbits were divided into control, model and intervention groups by random number table method, with 20 rabbits in each group.Prednisolone acetate (7.5 mg/kg) was injected bilaterally into the gluteal muscle of rabbits in both model and intervention groups twice a week.β-Ecd (0.5 mg/kg) was injected subcutaneously at the time of the first injection of Prednisolone acetate in the intervention group for 5 times a week.An equal amount of saline was injected into rabbits in control and model groups.Eight weeks after modelling, animals were put to death, and femoral heads were taken from both sides for gross observation.Micro computed tomography(Micro-CT) was used to analyze the microstructure of bone trabeculae and to measure bone microstructural parameters.Histological staining was used to detect changes in the morphology of bone tissues.Immunohistochemistry and Western blot were used to examine the expression of osteogenic and lipogenic factors and proteins in the femoral head tissue.Rabbit bone marrow mesenchymal stem cells (BMSCs) were divided into a blank control group, a Dexamethasone group and a β-Ecd intervention group.Cell damage was induced by Dexamethasone in Dexamethasone group and β-Ecd intervention group, and the β-Ecd intervention group was given the optimal concentration of β-Ecd.Western blot was used to detect the expression of osteogenic and lipogenic proteins in the cells of each group.Real-time quantitative polymerase chain reaction (RT-qPCR) was used to determine the mRNA expression of osteogenic and lipogenic marker genes.Results:After excluding 9 rabbits that died during the experimental period, 51 rabbits were finally included in the study, with 19 in the control group, 15 in the model group and 17 in the intervention group.Gross observation and Micro-CT showed that compared with that of the control group, the femoral head of the model group was obscure and greyish, with dark red necrotic areas.The bone trabeculae of the model group were sparse, thinned, disordered, and partially fractured, compared with those of the control group.The changes in the femoral head and bone trabeculae of the intervention group were between those of control and model groups.The bone mineral density, the number, thickness and relative volume of bone trabeculae significantly decreased and trabecular separation significantly increased in both model and intervention groups, compared with those in the control group ( F=12.78, 45.52, 32.74, 64.08, 8.83, all P<0.05).However, these symptoms in the intervention group were better than those in the model group.Pathological histology showed that in the control group, bone trabeculae were neatly arranged, robust and full, with a high number of osteoclasts and occasional empty bone sockets.In the model group, bone trabeculae were sparsely arranged and broken, with fewer osteoclasts, and the number of empty bone sockets increased and enlarged.In the intervention group, bone trabeculae had a more complete morphology, with fewer necrotic osteoclasts and reduced empty bone sockets, compared with the model group.Immunohistochemistry results showed that compared with the control group, the model group and intervention group had increased content of fatty acid binding proteins (FABP) and CCAAT/enhancer binding proteins α (CEBP) in the femoral head bone tissue, and decreased content of osteopontin (OPN) and Runt-related transcription factor 2 (RUNX2) .The changes in each index were greater in the model group than those in the intervention group ( F=21.07, 24.06, 17.92, 21.36, all P<0.05). Western blot detection showed that compared with the control group, the CEBP protein expression content of the femoral head in the model group and the intervention group was increased and the RUNX2 protein expression content was decreased. The changes of CEBP and RUNX2 were greater in the model group than those in the intervention group( F=73.43, 197.87, all P<0.05).Western blot detection of BMSCs showed that compared with the blank control group, the Dexamethasone group and β-Ecd intervention group had decreased expression of OPN and RUNX2 proteins and increased expression of FABP and CEBP proteins ( F=161.61, 358.01, 91.18, 69.04, all P<0.05).The changes in each index in the β-Ecd intervention group were smaller than those in the Dexamethasone group.RT-qPCR detection of BMSCs showed that in the Dexamethasone group had lower expression of OPN and RUNX2 and higher expression of CEBP and FABP than the blank control group ( F=19.71, 45.08, 61.46, 15.12, all P<0.05).The changes in each index in the β-Ecd intervention group were smaller than those in the Dexamethasone group. Conclusions:β-Ecd can attenuate the pathological changes of hormonal femoral head necrosis in young rabbits by promoting osteogenic differentiation of BMSCs, inhibiting lipogenic differentiation of BMSCs, and improving bone microstructure.

Objective:To investigate the effect of β-Ecdysterone (β-Ecd), an active ingredient in cow′s knee, on hormonal femoral head necrosis in young rabbits and explore the mechanism involved.Methods:An animal study.Sixty New Zealand young rabbits were divided into control, model and intervention groups by random number table method, with 20 rabbits in each group.Prednisolone acetate (7.5 mg/kg) was injected bilaterally into the gluteal muscle of rabbits in both model and intervention groups twice a week.β-Ecd (0.5 mg/kg) was injected subcutaneously at the time of the first injection of Prednisolone acetate in the intervention group for 5 times a week.An equal amount of saline was injected into rabbits in control and model groups.Eight weeks after modelling, animals were put to death, and femoral heads were taken from both sides for gross observation.Micro computed tomography(Micro-CT) was used to analyze the microstructure of bone trabeculae and to measure bone microstructural parameters.Histological staining was used to detect changes in the morphology of bone tissues.Immunohistochemistry and Western blot were used to examine the expression of osteogenic and lipogenic factors and proteins in the femoral head tissue.Rabbit bone marrow mesenchymal stem cells (BMSCs) were divided into a blank control group, a Dexamethasone group and a β-Ecd intervention group.Cell damage was induced by Dexamethasone in Dexamethasone group and β-Ecd intervention group, and the β-Ecd intervention group was given the optimal concentration of β-Ecd.Western blot was used to detect the expression of osteogenic and lipogenic proteins in the cells of each group.Real-time quantitative polymerase chain reaction (RT-qPCR) was used to determine the mRNA expression of osteogenic and lipogenic marker genes.Results:After excluding 9 rabbits that died during the experimental period, 51 rabbits were finally included in the study, with 19 in the control group, 15 in the model group and 17 in the intervention group.Gross observation and Micro-CT showed that compared with that of the control group, the femoral head of the model group was obscure and greyish, with dark red necrotic areas.The bone trabeculae of the model group were sparse, thinned, disordered, and partially fractured, compared with those of the control group.The changes in the femoral head and bone trabeculae of the intervention group were between those of control and model groups.The bone mineral density, the number, thickness and relative volume of bone trabeculae significantly decreased and trabecular separation significantly increased in both model and intervention groups, compared with those in the control group ( F=12.78, 45.52, 32.74, 64.08, 8.83, all P<0.05).However, these symptoms in the intervention group were better than those in the model group.Pathological histology showed that in the control group, bone trabeculae were neatly arranged, robust and full, with a high number of osteoclasts and occasional empty bone sockets.In the model group, bone trabeculae were sparsely arranged and broken, with fewer osteoclasts, and the number of empty bone sockets increased and enlarged.In the intervention group, bone trabeculae had a more complete morphology, with fewer necrotic osteoclasts and reduced empty bone sockets, compared with the model group.Immunohistochemistry results showed that compared with the control group, the model group and intervention group had increased content of fatty acid binding proteins (FABP) and CCAAT/enhancer binding proteins α (CEBP) in the femoral head bone tissue, and decreased content of osteopontin (OPN) and Runt-related transcription factor 2 (RUNX2) .The changes in each index were greater in the model group than those in the intervention group ( F=21.07, 24.06, 17.92, 21.36, all P<0.05). Western blot detection showed that compared with the control group, the CEBP protein expression content of the femoral head in the model group and the intervention group was increased and the RUNX2 protein expression content was decreased. The changes of CEBP and RUNX2 were greater in the model group than those in the intervention group( F=73.43, 197.87, all P<0.05).Western blot detection of BMSCs showed that compared with the blank control group, the Dexamethasone group and β-Ecd intervention group had decreased expression of OPN and RUNX2 proteins and increased expression of FABP and CEBP proteins ( F=161.61, 358.01, 91.18, 69.04, all P<0.05).The changes in each index in the β-Ecd intervention group were smaller than those in the Dexamethasone group.RT-qPCR detection of BMSCs showed that in the Dexamethasone group had lower expression of OPN and RUNX2 and higher expression of CEBP and FABP than the blank control group ( F=19.71, 45.08, 61.46, 15.12, all P<0.05).The changes in each index in the β-Ecd intervention group were smaller than those in the Dexamethasone group. Conclusions:β-Ecd can attenuate the pathological changes of hormonal femoral head necrosis in young rabbits by promoting osteogenic differentiation of BMSCs, inhibiting lipogenic differentiation of BMSCs, and improving bone microstructure.

Objective:To evaluate the efficacy and safety of a China original liquid pulsation system for the treatment of meibomian gland dysfunction (MGD).Methods:A non-randomized controlled clinical trial was conducted.Twenty-two patients (44 eyes) diagnosed with MGD in Eye and ENT Hospital of Fudan University from February to August 2022 were enrolled.The patients were assigned into two groups according to their willingness.Of the 22 patients (44 eyes), 10 patients (20 eyes) in single liquid pulsation system group were treated with single liquid pulsation system for 12 minutes, and 12 patients (24 eyes) in intense pulsed light (IPL) group were treated with a course (4 times) of IPL, warm compresses and meibomian gland massage at three-week intervals.There was no difference in age and other baseline clinical indexes between the two groups (all at P<0.05). The meibum grading, quality grading of tear film lipid layer, Symptom Assessment Questionnaire in Dry Eye (SANDE) questionnaire score, first and average tear breakup time (BUT), corneal fluorescein sodium staining (CFS) score, tear meniscus height (TMH), and the area of meibomian gland loss were determined at baseline, 1 and 3 months after treatment.This study adhered to the Declaration of Helsinki.The study protocol was approved by the Ethics Committee of Eye and ENT Hospital of Fudan University (No.2021069). Written informed consent was obtained from each patient before any medical examination. Results:Statistically significant group effects and time effects were found in the quality of tear film lipid layer ( Hgroup=4.39, P=0.036, Htime=6.30, P=0.043) and average BUT ( Fgroup=4.41, P=0.038; Ftime=4.08, P=0.049) in the two groups.The meibum grading, first BUT and TMH 1 and 3 months after treatment were significantly better than before treatment in single liquid pulsation system group (all at P<0.05). Compared with before treatment, there was no significant improvement in the meibum grading, distribution of tear film lipid, first BUT and TMH at 1 and 3 months after treatment in IPL group (all at P>0.05). In both groups, the SANDE and CFS scores 1 and 3 months after treatment were better than those before treatment, showing statistically significance (all at P<0.05). In terms of safety, neither instrument-related adverse events nor extra complaints of discomfort were reported in the single liquid pulsation system group.In both groups, the number of patients with positive CFS staining significantly decreased, and no new cases with positive CFS appeared after treatment. Conclusions:This China original liquid pulsation system is a safe and effective physical therapy in improving tear film dysfunction and ocular surface symptoms of MGD patients within 3 months after treatment.

Objective: To assess clinical features and treatment outcomes in immunocompetent patients with primary central nervous system lymphoma (PCNSL). Methods: Sixty-two patients with PCNSL who attended Guangdong General Hospital between January 1998 and January 2012 were included. Survival curves were estimated using Kaplan-Meier survival methodology and statistical significance of continuous was assessed via the Cox proportional hazard model. Results: The median age of the patient cohort was 56 years, and the male to female ratio was 1.14∶1.00. The common presentations were increased intracranial pressure symptoms and neuron damage. Performance status of 54 (54/62, 87.1%) patients were the international prognostic index (IPI) 0-2. Diffuse large B-cell lymphoma (57/62, 91.9%) was most common, and the rest were T-cell lymphoma (4/62,6.4%) and extranodal marginal zone lymphoma of mucosa-associated lymphoid tissue (1/62, 1.6%). In the series, 32 patients (32/62, 51.6%) had multiple lesions. Involvement of deep structures was found in 30 (30/62, 48.4%) patients. An elevated serum LDH level was detected in 19 (19/62, 30.6%) patients and the Ki-67 index was ≥90% in 38 (38/62, 61.3%) patients. Univariate analysis showed patients who were female, age<60 years, had WHO Eastern Cooperative Oncology Group performance status grade 0-2, single lesion, absence of deep structures involvement and normal LDH level showed better 2-year survival rate and longer median survival time. Significance was only seen in the normal LDH level group. Multivariate Cox regression analysis revealed that radical surgery only and Rituximab+ high-dose of methotrexate+ whole brain radiation therapy (WBRT) were independent prognostic indicators in PCNSL patients (P<0.05). Conclusions PCNSL is a rare but aggressive tumor with poor prognosis. Patients treated with high-dose of methotrexate combining with rituximab, followed by WBRT have a better prognosis and longer survival time, and thus these could probably be a promising treatment.

assess clinical features and treatment outcomes in immunocompetent patients with primary central nervous system lymphoma (PCNSL). Methods Sixty?two patients with PCNSL who attended Guangdong General Hospital between January 1998 and January 2012 were included. Survival curves were estimated using Kaplan?Meier survival methodology and statistical significance of continuous was assessed via the Cox proportional hazard model. Results The median age of the patient cohort was 56 years, and the male to female ratio was 1.14∶1.00. The common presentations were increased intracranial pressure symptoms and neuron damage. Performance status of 54 (54/62, 87.1%) patients were the international prognostic index (IPI) 0-2. Diffuse large B?cell lymphoma (57/62, 91.9%) was most common, and the rest were T?cell lymphoma (4/62, 6.4%) and extranodal marginal zone lymphoma of mucosa?associated lymphoid tissue (1/62, 1.6%). In the series, 32 patients (32/62, 51.6%) had multiple lesions. Involvement of deep structures was found in 30 (30/62, 48.4%) patients. An elevated serum LDH level was detected in 19 (19/62, 30.6%) patients and the Ki?67 index was ≥90% in 38 (38/62, 61.3%) patients. Univariate analysis showed patients who were female, age<60 years, had WHO Eastern Cooperative Oncology Group performance status grade 0-2, single lesion, absence of deep structures involvement and normal LDH level showed better 2?year survival rate and longer median survival time. Significance was only seen in the normal LDH level group. Multivariate Cox regression analysis revealed that radical surgery only and Rituximab+ high?dose of methotrexate+ whole brain radiation therapy (WBRT) were independent prognostic indicators in PCNSL patients (P<0.05). Conclusions PCNSL is a rare but aggressive tumor with poor prognosis. Patients treated with high?dose of methotrexate combining with rituximab, followed by WBRT have a better prognosis and longer survival time, and thus these could probably be a promising treatment.

OBJECTIVETo construct a lentiviral expression vector for human VHL and its shRNA vector, and study the effect of VHL on proliferation and apoptosis of renal cell carcinoma cell lines.

METHODSLentiviral vectors pZsGreen1-VHL and pLL3.7-shVHL were constructed and transfected into 293T cells with 3 packaging plasmids by Lipofectamine(TM) 2000 reagent. The supernatant was collected to infect A498 and Caki-1 cells, respectively. VHL mRNA and protein levels were detected by RT-PCR and Western blotting, respectively. The effect of VHL on the proliferation, cell cycle and cell apoptosis were analyzed by MTS and flow cytometry.

RESULTSThe recombinant lentiviral vectors were successfully constructed. The proliferation of A498 cells with reconstituted wild-type VHL was significantly inhibited, while the proliferation of Caki-1 cells with VHL knockdown was significantly enhanced as compared with the control cells (P<0.05). VHL induced G0/G1-S cell cycle arrest. The apoptosis rate of A498 cells with reconstituted wild-type VHL was significantly increased while that of Caki-1 cells with VHL knockdown was significantly lowered compared with the control cells (P<0.05).

CONCLUSIONVHL can inhibit the proliferation and induce apoptosis of renal cell carcinoma cells.

Apoptosis ; Carcinoma, Renal Cell ; pathology ; Cell Cycle ; Cell Line, Tumor ; Cell Proliferation ; Genetic Vectors ; Humans ; Lentivirus ; Plasmids ; RNA, Messenger ; RNA, Small Interfering ; Transfection ; Von Hippel-Lindau Tumor Suppressor Protein ; genetics

Objective To construct a lentiviral expression vector for human VHL and its shRNA vector, and study the effect of VHL on proliferation and apoptosis of renal cell carcinoma cell lines. Methods Lentiviral vectors pZsGreen1-VHL and pLL3.7-shVHL were constructed and transfected into 293T cells with 3 packaging plasmids by LipofectamineTM 2000 reagent. The supernatant was collected to infect A498 and Caki-1 cells, respectively. VHL mRNA and protein levels were detected by RT-PCR and Western blotting, respectively. The effect of VHL on the proliferation, cell cycle and cell apoptosis were analyzed by MTS and flow cytometry. Results The recombinant lentiviral vectors were successfully constructed. The proliferation of A498 cells with reconstituted wild-type VHL was significantly inhibited, while the proliferation of Caki-1 cells with VHL knockdown was significantly enhanced as compared with the control cells (P<0.05). VHL induced G0/G1-S cell cycle arrest. The apoptosis rate of A498 cells with reconstituted wild-type VHL was significantly increased while that of Caki-1 cells with VHL knockdown was significantly lowered compared with the control cells (P<0.05). Conclusion VHL can inhibit the proliferation and induce apoptosis of renal cell carcinoma cells.

Objective To construct a lentiviral expression vector for human VHL and its shRNA vector, and study the effect of VHL on proliferation and apoptosis of renal cell carcinoma cell lines. Methods Lentiviral vectors pZsGreen1-VHL and pLL3.7-shVHL were constructed and transfected into 293T cells with 3 packaging plasmids by LipofectamineTM 2000 reagent. The supernatant was collected to infect A498 and Caki-1 cells, respectively. VHL mRNA and protein levels were detected by RT-PCR and Western blotting, respectively. The effect of VHL on the proliferation, cell cycle and cell apoptosis were analyzed by MTS and flow cytometry. Results The recombinant lentiviral vectors were successfully constructed. The proliferation of A498 cells with reconstituted wild-type VHL was significantly inhibited, while the proliferation of Caki-1 cells with VHL knockdown was significantly enhanced as compared with the control cells (P<0.05). VHL induced G0/G1-S cell cycle arrest. The apoptosis rate of A498 cells with reconstituted wild-type VHL was significantly increased while that of Caki-1 cells with VHL knockdown was significantly lowered compared with the control cells (P<0.05). Conclusion VHL can inhibit the proliferation and induce apoptosis of renal cell carcinoma cells.

The interaction mechanism between rhaponticin (RT) and human serum albumin (HSA) has been studied by fluorescence spectroscopy and absorbance spectra. The mediation effect that the metal ions took part in the interaction has also been discussed in this paper. Based on different theoretical models of fluorescence quenching, the binding constant (K) and binding sites (n) of the interaction were determined and analyzed comparatively. The quenching mechanism of the binding reaction has also been discussed. The binding distance (r) and energy-transfer efficiency (E) between RT/RT-Co(II)/RT-Ni(II) and HSA were also obtained by virtue of the Förster theory of non-radiation energy transfer. The effect of RT acting on the HSA's conformation was analyzed by synchronous fluorescence spectroscopy. The result showed that the result calculated by different theoretical models is generally equivalent and RT bound HSA strongly by forming stable complex, which indicates that HSA under physiological conditions can act as a carrier for RT to be transported to exert effects. The microconformation of HSA changed significantly due to hydrophobicity change in the chemical environment of some fluorescence chromophores in the subdomain IIA and IIB of HSA. Metal ions Co(II) and Ni(II) can mediate RT-HSA interaction, making the binding of the drug to protein stronger, which indicates that Co(II) and Ni(II) can enhance rhaponticin's medical efficacy under physiological conditions.

Objective To observe the clinical curative effect of treating CHD angina with ShenQiHuaYu decoction. Methods 100 patients with angina were recruited into a control group and a treatment group, with 50 patients in each group. The treatment group was treated with ShenQiHuaYu diction, and the control group was treated with conventional western medicine. Results The total effective rate and total effective rate for ECG in the treatment group and the control group was 91.6%, 81.2%, and 77%, 67%. There was significant difference between the two group (P<0.05). Conclusion ShenQiHua Yu decoction is effective in treating CHD angina.