Mechanical Stimulus Inhibits the Growth of a Bone Tissue Model Cultured In Vitro

Objectives To construct the cancellous bone explant model and a method of culturing these bone tissues in vitro,and to investigate the effect of mechanical load on growth of cancellous bone tissue in vitro.Methods Cancellous bone were extracted from rabbit femoral head and cut into 1-mm-thick and8-mm-diameter slices under sterile conditions.HE staining and scanning electron microscopy were employed to identify the histomorphology of the model after being cultured with a new dynamic load and circulating perfusion bioreactor system for 0,3,5,and 7 days,respectively.We built a three-dimensional model using microCT and analyzed the loading effects using finite element analysis.The model was subjected to mechanical load of 1000,2000,3000,and 4000μεrespectively for 30 minutes per day.After 5 days of continuous stimuli,the activities of alkaline phosphatase(AKP)and tartrate-resistant acid phosphatase(TRAP)were detected.Apoptosis was analyzed by DNA ladder detection and caspase-3/8/9 activity detection.Results After being cultured for 3,5,and 7 days,the bone explant model grew well.HE staining showed the apparent nucleus in cells at the each indicated time,and electron microscope revealed the living cells in the bone tissue.The activities of AKP and TRAP in the bone explant model under mechanical load of 3000 and 4000μεwere significantly lower than those in the unstressed bone tissues(all P<0.05).DNA ladders were seen in the bone tissue under 3000 and 4000μεmechanical load.Moreover,there was significant enhancement in the activities of caspase-3/8/9 in the mechanical stress group of 3000 and 4000με(all P<0.05).Conclusions The cancellous bone explant model extracted from the rabbit femoral head could be alive at least for 7 days in the dynamic load and circulating perfusion bioreactor system,however,pathological mechanical load could affect the bone tissue growth by apoptosis in vitro.The differentiation of osteoblasts and osteoclasts might be inhibited after the model is stimulated by mechanical load of 3000 and 4000με.

The efficacy of TKIs in treatment of human primary small cell lung cancer xenograft model in vivo

Objective: To study the treatmaient of non-small cell lung cancer, we established the HU-Prim allograft transplantation tumor model. Methods: The fresh tumor samples were transplanted in the right scapular subcutaneous layer of the severe combined immunodeficient Non-obese diabetic/severe combined immunodeficient(NOD/SCID) mice. The pathological features of the tumors were observed. Nonnecrotic tissue was inoculated subcutaneously into the right axillary. When the tumor in burdened rat grew approximately 100 mm3, according to the tumor size all the animals were divided into the following four groups, eight rats in each group: solvent control group, gefitinib group(100 mg/kg), erlotinib group(50 mg/kg), afatinib group(20 mg/kg). Aniamals were treated with drugs by intragastric(i.g.) administrated, once daily, for consecutively 14 days. Measure the tumor size 2-3 times every week. Results: Hu Prime1-NSCLC mutant sensitive xenograft model research data showed that reversible tyrosine kinase inhibitors gefitinib, erlotinib and irreversible tyrosine kinase inhibitor afatinib could effectively inhibit tumor growth in EGFR positive NSCLC allografts model. The pharmacodynamic activity of irreversible inhibitor was better than that of the reversible inhibitor. Specimens from clinical anthropogenic tumor retain characteristics of the human primary malignancy, histopathology, biological characteristics, and tumor markers, etc., which can more accurately reflect the characteristics of the tumor and the impact of interventions. Conclusion: The model is not only a good antitumor drug experimental platform, but also a new evaluation tool of individualized medication.

An uncontrolled open-label, multicenter study to monitor the antiviral activity and safety of inhaled zanamivir （as Rotadisk via Diskhaler device） among Chinese adolescents and adults with influenza-like illness

Background It is the first multicenter clinical study in China to investigate zanamivir use among Chinese adolescents and adults with influenza-like illness （ILl） since 2009, when inhaled zanamivir （RELENZA ） was marketed in China. Methods An uncontrolled open-label, multicentre study to evaluate the antiviral activity, and safety of inhaled zanamivir （as Rotadisk via Diskhaler device）; 10 mg administered twice daily for 5 days in subjects≥12 years old with ILl. Patients were enrolled within 48 hours of onset and followed for eight days. Patients were defined as being influenza-positive if the real-time reverse transcriptase-polymerase chain reaction real-time reverse transcriptase-polymerase chain reaction （rRT-PCR） test had positive results. Results A total of 400 patients 〉12 years old were screened from 11 centers in seven provinces from March 2010 to January 2011. Three hundred and ninety-two patients who took at least one dose of zanamivir were entered into the safety analysis. The mean age was 33.8 years and 50% were male. Cardiovascular diseases and diabetes were the most common comorbidities. All the reported adverse events, such as rash, nasal ache, muscle ache, nausea, diarrhea, headache, occurred in less than 1% of subjects. Mild sinus bradycadia or arrhythmia occurred in four subjects （1%）. Most of the adverse events were mild and did not require any change of treatment. No severe adverse events （SAE） or fatal cases were reported. Bronchospasm was found in a 38 years old woman whose symptoms disappeared after stopping zanamivir and without additional treatment. All the 61 influenza virus isolates （43 before enrollment, 18 during treatment） proved to be sensitive to zanamivir. Conclusions Zanamivir is well tolerated by Chinese adolescents and adults with ILls. There is no evidence for the emergence of drug-resistant isolates during treatment with zanamivir. （ChiCTR-TNRC-10000938）

Effect of Puerarin on the Release of lnterleukin-8 in Co-Culture of Human Bronchial Epithelial Cells and Neutrophils

Objective： To investigate the effect of puerarin on intedeukin （IL）-8 mRNA expression and the protein release in the co-culture of human bronchial epithelial （BEAS-2B） cells and human neutrophils. Methods： BEAS- 2B cells and neutrophills were cultured separately and co-cultured with puerarin （50, 100, and 200 ug/mL） for a predetermined time. Cytokines in culture supematant were evaluated by protein array and IL-8 quantified by enzymelinked immunosorbent assay （ELISA）. IL-8 mRNA expression was evaluated by real-time quantitative polymerase chain reaction （real-time qPCR）. Results： The co-culture of BEAS-2B cells and neutrophils exhibited synergistic effects on IL-8 mRNA expression in BEAS-2B cells, but not in neutrophils after 12 h incubation （P〈0.01）, as compared with that in BEAS-2B cells or neutrophils alone. IL-8 protein release in the culture supernatant was obviously elevated when BEAS-2B cells were co-cultured with human neutrophils as compared with that in the supematant of BEAS-2B cells or neutrophils alone after incubated for 2, 6, 12, and 18 h （P〈0.01）. Treatment with puerarin could significantly down-regulate the expression of IL-8 mRNA in BEAS-2B cells and IL-8 release in the supematant of the co-culture of BEAS-2B cells and neutrophils （P〈0.01）. Conclusion： Puerarin could exhibit anti-inflammatory activity by suppressing IL-8 production from the co-culture of human bronchial epithelial cells and neutrophils.

Potent anti-angiogenesis and anti-tumor activity of a nove human anti-VEGF antibody, MIL60

Angiogenesis is crucial for tumor development, growth and metastasis. Vascular endothelial growth factor （VEGF） has been implicated in promoting solid tumor growth and metastasis via stimulating tumor-associated angiogenesis, and blocking the activity of VEGF can starve tumors. Avastin, which is a humanized anti-VEGF antibody, has been successfully applied in clinics since 2004. However, the price of Avastin is extremely high for Chinese people. Here, we report a novel human anti-VEGF neutralizing antibody, MIL60, which shows an affinity comparable to that of Avastin （the KD value of MIL60 was 44.5 pM, while that of Avastin was 42.7 pM）. MIL60 displays favorable actions in inhibiting VEGF-triggered endothelial cell proliferation （the IC5o value of M IL60 was 31-6.4 ng/ml and that of Avastin was 47--.8.1 ng/ml）, migration （8 pg/ml or 0.8 pg/ml MIL60 versusthe control： P〈O.05） and tube formation （2 I^g/ml or 0.2 lzg/ml MIL60 versusthe control： P〈O.05） viathe VEGFR2 signaling pathway. Moreover, MIL60 was shown to inhibit tumor growth and angiogenesis in vivo in xenograft models of human colon carcinoma and ovarian cancer using immunotherapy and immunohistochemistry analysis （MIL60 versus N.S.： P=0.0007; Avastin versus N.S.： P=0.00046）. These data suggest that MIL60 is a potential therapeutic, anti-angiogenic agent. Our work provides a novel anti-VEGF antibody, which can be considered an anti-tumor antibody candidate and a new option for patients with various cancers.

Epidemiology and evolution of Middle East respiratory syndrome coronavirus, 2012-2020

Background:The ongoing transmission of the Middle East respiratory syndrome coronavirus(MERS-CoV)in the Middle East and its expansion to other regions are raising concerns of a potential pandemic.An in-depth analysis about both population and molecular epidemiology of this pathogen is needed.Methods:MERS cases reported globally as of June 2020 were collected mainly from World Health Organization official reports,supplemented by other reliable sources.Determinants for case fatality and spatial diffusion of MERS were assessed with Logistic regressions and Cox proportional hazard models,respectively.Phylogenetic and phylogeographic analyses were performed to examine the evolution and migration history of MERS-CoV.Results:A total of 2562 confirmed MERS cases with 150 case clusters were reported with a case fatality rate of 32.7%(95%Cl:30.9-34.6%).Saudi Arabia accounted for 83.6%of the cases.Age of>65 years old,underlying conditions and>5 days delay in diagnosis were independent risk factors for death.However,a history of animal contact was associated with a higher risk(adjusted OR=297,95%Cl:1」0-7.98)among female cases<65 years but with a lower risk(adjusted OR=0.31,95%Cl:0.18-0.51)among male cases>65 years old.Diffusion of the disease was fastest from its origin in Saudi Arabia to the east,and was primarily driven by the transportation network.The most recent subclade C5.1(since 2013)was associated with non-synonymous mutations and a higher mortality rate.Phylogeographic analyses pointed to Riyadh of Saudi Arabia and Abu Dhabi of the United Arab Emirates as the hubs for both local and international spread of MERS-CoV.Conclusions:MERS-CoV remains primarily locally transmitted in the Middle East,with opportunistic exportation to other continents and a potential of causing transmission clusters of human cases.Animal contact is associated with a higher risk of death,but the association differs by age and sex.Transportation network is the leading driver for the spatial diffusion ofthe disease.These findings how this pathogen spread are helpful for targeting public health surveillance and interventions to control endemics and to prevent a potential pandemic.

Five new benzophenone glycosides from the leaves of Aquilaria sinensis(Lour.) Gilg

Phytochemical investigation on the ethanol extract from the leaves of Aquitaria sinensis led to the isolation of five new benzophenone glycosides,aquilarinensides A-E（1-5）.Their structures were elucidated by a combination of 1D and 2D NMR,HRMS,and chemical analysis.