First screening for tick-borne pathogens in Chinese Milu deer (Elaphurus davidianus).

Ticks are primary vectors for many tick-borne pathogens (TBPs) and pose a serious threat to veterinary and public health. Information on the presence of TBPs in Chinese Milu deer (Elaphurus davidianus) is limited. In this study, a total of 102 Chinese Milu deer blood samples were examined for Anaplasma spp., Theileria spp., Babesia spp., Rickettsia spp., and Borrelia spp., and three TBPs were identified: Anaplasma phagocytophilum (48; 47.1 %), Candidatus Anaplasma boleense (47; 46.1%), and Theileria capreoli (8; 7.8 %). Genetic and phylogenetic analysis of the 16S rRNA and 18S rRNA confirmed their identity with corresponding TBPs. To our knowledge, this is the first report on Candidatus A. boleense and T. capreoli detection in Chinese Milu deer. A high prevalence of A. phagocytophilum with veterinary and medical significance was identified in endangered Chinese Milu deer, which could act as potential zoonotic reservoirs. The identification of the TBPs in Chinese Milu deer provides useful information for the prevention and control of tick-borne diseases.

[Textual and information research of classical prescription Mijiao Pills].

Elaphurus davidianus( Milu),a rare animal unique to China,has been used as medicine for more than a thousand years,but the extinction of Milu in modern times resulted in the unavailability of related medical products. Today,the reintroduction of Milu population makes it possible to restore its medicinal usage. The resource reserves of Cervi Cornu,the natural shedding product from Milu,are increasing with the expansion of the population,allowing it to be fully utilized in the medical field. Mijiao Pills,first recorded in Important Prescriptions Worth a Thousand Gold for Emergency( Bei Ji Qian Jin Yao Fang) by Sun Simiao in the Tang Dynasty,is the first Chinese medicinal prescription with Cervi Cornu as the sovereign medicinal,which is effective in tonifying. Its composition,preparation,efficacy and indications,and administration are described in detail in the Important Prescriptions Worth a Thousand Gold for Emergency,which however,have changed significantly over the thousands of years,seriously affecting the clinical application of this classical prescription and related product development. Therefore,the key information of this prescription should be systematically collated and summarized. According to the principles of textual research on key information in ancient classical prescriptions promulgated by relevant authorities,this paper reviewed ancient Chinese medical books of the past dynasties,modern literature,as well as the Pharmacopoeia of the People's Republic of China( 2020 Version) to figure out such key information as the source,historical evolution,original plants and animals and their processing,dosage,preparation,and usage of Mijiao Pills. This paper aimed to provide a basis for the clinical application of Mijiao Pills and subsequent product development,thus facilitating the development and utilization of this precious medicinal animal resource.

[Passivation of Simulated Pb-and Cd-Contaminated Soil by Applying Combined Treatment of Phosphate, Humic Acid, and Fly Ash].

In this study, three kinds of amendments including superphosphate, humic acid, and fly ash and their complex combination were adopted to passivate the artificially simulated Pb-and Cd-containing soils. The passivation efficiency evaluation was performed via the CaCl2 and triethylenetriaminepentaacetic acid (DTPA) extraction method as well as a BCR morphological classification experiment. The microstructures and structures of the soil were explored further via X-ray diffraction (XRD) and scanning electron microscopy with X-ray energy dispersive spectroscopy (SEM-EDS) to elaborate the passivation mechanism. The results demonstrated that all passivation processes, excluding single humic acid addition, could reduce the CaCl2 and DTPA extraction contents of Pb and Cd in soils, where the optimal efficiency could be achieved by the sequential addition of superphosphate and humic acid, followed by fly ash. There was a weakly positive correlation between soil pH and CaCl2/DTPA extraction content of Pb, a negative correlation between soil pH and CaCl2/DTPA extraction content of Cd, and a significantly negative correlation between available phosphorous content and CaCl2/DTPA extraction contents of Pb and Cd, suggesting the crucial role of available phosphorous contents to control the activities of Pb and Cd. In the presence of phosphate, humic acid, and fly ash, the Pb and Cd could convert from active weak acid extraction to low-activity residual speciation, resulting in effectively reducing Pb and Cd transferability. Throughout the XRD and SEM-EDS analyses, it was found that ion exchange was the predominant mechanism in heavy metal passivation by single superphosphate, wherein the heavy metals were transformed into an insoluble Ca-containing phosphate mixture. The dissolving/precipitation or surface adsorption could be concluded as the main mechanism in the combination of the three passivation agents that converted heavy metals to lead phosphate precipitate[(Pb3(PO4)2] or mixed heavy metal mineral[PbFe3(SO4)(PO4)(OH)6], so as to obtain superior heavy metal passivation achievement.

Effect of metformin on the proliferation and apoptosis of the renal cancer cell line 786-O and the underlying mechanisms.

PURPOSE: To investigate the effects of metformin (Met) on the proliferation and apoptosis of a renal carcinoma cell line and study the underlying mechanisms. METHODS: Renal cell carcinoma (RCC) line 786-O was cultured with media containing different concentrations of Met. The proliferation and apoptosis of 786-O cells were detected by the MTT method and flow cytometry, respectively. The invasion of 786-O cells was detected by scratch test, and the expression of pro-apoptotic proteins was determined by Western blot assay. RESULTS: The proliferation rates of 786-O cells with Met concentrations of 10, 15, and 20 mM were decreased by 62.8, 61.7, and 65.1%, respectively, with no significant difference among these concentrations (p>0.05). Twenty-four hrs after the scratch assay, the mean migration index in the control group and Met treatment group was 51.6% +/- 5.9 and 28.1% +/- 4.3, and that after 48 hrs was 92.2% +/- 6.4 and 68.0% +/- 4.9, respectively (p<0.05). At low serum concentration, the percentage of apoptotic cells in the Met treatment group (17.6%) was significantly higher (p<0.05) than that in the control group (5.2%). Met (10 mM) treatment significantly increased the expression of Caspase-3 and Bax proteins in 786-O cells (p<0.05). CONCLUSIONS: Metformin may be a potential drug of choice in the treatment of metastatic RCC.

[Stabilization Treatment of Pb and Zn in Contaminated Soils and Mechanism Studies].

In the present work, the combined application of potassium dihydrogen phosphate, quick lime and potassium chloride was used to immobilize the Pb and Zn in contaminated soils. The efficiency of the process was evaluated through leaching tests and Tessier sequential extraction procedure. The mechanism of stabilization was analyzed by X-ray diffraction (XRD) and scanning electron microscope (SEM) to reveal the mechanism of stabilization. The results showed that the stabilizing efficiency of Pb contaminated soils was above 80% and the leaching concentrations of Pb, Zn were far below the threshold when the ratio of exogenous P and soil (mol · mol⁻¹) was 2:1-4: 1, the dosing ratio of CaO was 0.1%-0.5% ( mass fraction) and the dosage of potassium chloride was 0.02-0. 04 mol. Meanwhile, Pb and Zn in soil were transformed from the exchangeable fraction into residual fraction, which implied that the migration of Pb, Zn in soil could be confined by the stabilization treatment. XRD and SEM analysis revealed that Ca-P-Pb precipitation, lead orthophosphate [PbHP04, Pb3 (PO4)2], pyromorphite (Pb-PO4-Cl/OH) and mixed heavy metal deposits (Fe-PO4- Ca-Pb-Zn-OH) could be formed after solidification/stabilization in which Pb and Zn could be wrapped up to form a solidified composition and to prevent leaching.

Efficacy and safety of the treatment: combination of benazepril/lercanidipine vs. benazepril alone in patients with mild-to-moderate hypertension.

BACKGROUND: Combination therapy is an effective method to reduce the blood pressure (BP) for patients with hypertension. This study was performed to evaluate the efficacy and safety of benazepril/lercanidipine compared with benazepril alone in patients with mild-to-moderate hypertension. METHODS: One hundred and eighty-one patients with mild-to-moderate primary hypertension were assigned in this randomized, single-blind, parallel-group study and were randomly divided into group A (benazepril 10 mg/lercanidipine 10 mg) and group B (benazepril 10 mg) for 8 weeks. At 4 weeks, the dosage of Benazepril was titrated up to 20 mg if the diastolic blood pressure (DBP) remained ≥ 90 mmHg. BP control and side effects were evaluated at the end of 1, 4 and 8 weeks. RESULTS: The baseline characteristics of the two groups were similar. The BP in both groups decreased from the baseline (P < 0.05). At the end of 4 and 8 weeks, Benazepril/Lercanidipine produced greater BP reduction than Benazepril alone (P < 0.05). The comparison of the rate of BP control for the benazepril/lercanidipine and benazepril groups at the end of 1, 4, and 8 weeks were 41.2% vs. 37.6% (P > 0.05), 67.1% vs. 44.7% (P < 0.05), and 71.8% vs. 45.9% (P < 0.05). There was no significant difference of side effects between the two groups. CONCLUSION: The benazepril/lercanidipine combination is more effective in reducing BP than benazepril alone, while it does not increase the incidence of side effects.

[Transfection of GFP mRNA in dendritic cells and analysis of some factors involved].

AIM: To investigate the transfection of green fluorescent protein (GFP) mRNA in dendritic cells (DC) and analyze some factors which influence the transfection efficiency. METHODS: GFP (as a report gene) mRNA with cap was synthesized, in vitro, with mMESSAGE RNA Transcription Kit containing T7 RNA polymerase, and then the poly(A) was added to the GFP caped-mRNA by yeast poly(A) polymerase. DC were generated from the monocytes isolated from human peripheral blood by stimulation of GM-CSF and IL-4. The GFP mRNA was transfected into DC mediated by transfection reagent. The transfection efficiency and the expression levels were measured by flow cytometry. RESULTS: GFP expression in DC has been obtained by transfecting its mRNA synthesized in vitro. The transfection reagents, mRNA concentrations and cell densities have the significant effects on the transfection. The high level of transfection efficiency (up to 27%) was obtained using Transmessenger Transfection Kit with 1 microg gfp mRNA in 200 microL X-VIVO-15 serum-free medium at the cell density of 2.5x10(9)/L. CONCLUSION: The high-level transfection efficiency of gfp gene in DC could been achieved by using GFP mRNA in the optimum transfection conditions.

High expression level of soluble SARS spike protein mediated by adenovirus in HEK293 cells.

AIM: To develop a highly efficacious method for preparation of soluble SARS S-protein using adenovirus vector to meet the requirement for S-protein investigation. METHODS: The human adenovirus vector was used to express the soluble S-protein (corresponding to 1 approximately 1190 amino acids) fused with Myc/His tag using codon-optimized gene construct in HEK239 cells. The recombinant adenovirus bearing S-protein gene was generated by ligation method. The expressed S-protein with Myc/His tag was purified from culture medium with Ni-NTA agarose beads followed by dialysis. The S-protein was detected by Western blot and its biologic activity was analyzed by binding to Vero cells. RESULTS: Under the conditions of infection dose (MOI of 50) and expression time (48 h), the high-level expression of S-protein was obtained. The expression level was determined to be approximately 75 microg/10(6) cells after purification. Purified soluble S-protein was readily detected by Western blot with anti-Myc antibody and showed the ability to bind to surface of Vero cells,demonstrating that the soluble S-protein could remain the biologic activity in the native molecule. CONCLUSION: The high-level expression of S-protein in HEK293 cells mediated by adenovirus can be achieved under the optimized expression conditions. The proteins possess the biologic activity, which lays a foundation for further investigation of S-protein biological function.

[Genetic mutations of homocysteine metabolism related enzymes in patients with ischemic stroke].

To study genetic mutations of methylenetetrahydrofolate reductase (MTHFR) C677T and cystathionine-beta-synthase (CBS) T833C related to homocysteine metabolism in patients with ischemic stroke, the MTHFR gene C677T gene mutation and the CBS T833C gene mutation were detected by PCR-RFLP or ARMS method in 74 patients with ischemic stroke and 83 normal people for control. Results showed that the frequencies of MTHFR T homogenetic type (2.7%) , heterogenetic type (51.4%) and T allele (28.4%) in ischemic group were higher than those in control group (1.2%, 39.8% and 21.1%, respectively). The frequencies of CBS C homogenetic type (13.5%) and C allele (43.9%) in ischemic group were higher than those in control group (6.0% and 38.0%, respectively). Multiple Logistic Regression analysis showed that together with the T allele in MTHFR, the C allele in CBS and age were related to ischemic stroke (P<0.05). The odds ratios (OR) of the T allele in MTHFR C677T and the C allele in CBS T833C were 1.74 (95%CI 1.06-2.86) and 1.73 (95%CI 1.07-2.81) respectively. The study revealed that the genetic mutations of MTHFR C677T, CBS T833C,were related with the ischemic stroke. The genetic mutations of MTHFR C677T and CBS T833C may be genetic factors for ischemic stroke.

[Cloning and expression of a gene encoding shortened cecropin A-melittin hybrid in E.coli].

According to the studies of David Andreu and other scientists, we designed and synthesized a gene encoding shortened cecropin A-melittin hybrid. This hybrid consists of CA(1-7) and M(5-12). The gene was inserted into pGEMEX-1 in the site of EcoRI and BamHI. A positive clone was obtained by hybridization and DNA sequencing and was expressed in JM109(DE3).