Progress in SARS-CoV-2, diagnostic and clinical treatment of COVID-19.

Background: Corona Virus Disease 2019(COVID-19)is a global pandemic novel coronavirus infection disease caused by Severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2). Although rapid, large-scale testing plays an important role in patient management and slowing the spread of the disease. However, there has been no good and widely used drug treatment for infection and transmission of SARS-CoV-2. Key findings: Therefore, this review updates the body of knowledge on viral structure, infection routes, detection methods, and clinical treatment, with the aim of responding to the large-section caused by SARS-CoV-2. This paper focuses on the structure of SARS-CoV-2 viral protease, RNA polymerase, serine protease and main proteinase-like protease as well as targeted antiviral drugs. Conclusion: In vitro or clinical trials have been carried out to provide deeper thinking for the pathogenesis, clinical diagnosis, vaccine development and treatment of SARS-CoV-2.

An acute gastroenteritis outbreak caused by GII.P16-GII.2 norovirus associated with airborne transmission via the air conditioning unit in a kindergarten in Lianyungang, China.

Noroviruses are a common cause of acute gastroenteritis outbreaks in institutions including schools and kindergartens around the world. An outbreak caused by GII.P16-GII.2 norovirus in a kindergarten in Lianyungang, Jiangsu Province, China is reported here. An epidemiological investigation was conducted, and pathogen detection was performed. The descriptive analysis indicated that this outbreak in middle class 1 had a point source. Twenty cases of acute gastroenteritis occurred in this class within a period of 8.5h; the attack rate was 52.6% (20/38). Airborne transmission via the air conditioning unit in a confined restroom could have played a critical role in this outbreak. Sequence analysis of GII-positive samples confirmed that the norovirus GII.P16-GII.2 variant was the etiological agent of this outbreak.

Protective role of berberine and Coptischinensis extract on T2MD rats and associated islet Rin­5f cells.

The aim of the present study was to compare the different effects of berberine (Ber) and Coptischinensis extract (CCE) on a rat model of type 2 diabetes mellitus (T2DM), and the islet Rin­5f cell line was used to examine the differences between Ber and CCE and the underlying mechanisms. CCE was extracted and purified prior to analysis. Male Sprague­Dawley rats were provided with a high­fat diet to induce insulin resistance prior to injecting with streptozotocinto establish the T2DM model, the T2DM rats were treated with Ber and CCE, and blood samples and pancreatic tissues were obtained and compared to examine T2DM metabolic syndromes among the groups of rats, which included healthy rats, model rats, and model rats treated with Ber and CCE at different doses between 0 and 8 weeks. The protective effects of Ber and CCE on the Rin­5f islet cell line were also evaluated. The effects on Rin­5f cell proliferation and cell cycle, glucose­stimulated insulin release test (GSIS), the anti­apoptotic effects caused by fat induction, and protein expression levels of poly ADP­ribose polymerase (PARP­1) were evaluated. The results showed that the content of the prepared CCE was 96.07% for five alkaloids. When it was used for treatment of the T2DM rats, compared with Ber, metformin and rosiglitazone, the fasting blood glucose, glucosylated serum protein (GSP) and glucose infusion rate indicesin the fasting rats were ameliorated, compared with those in the T2MD rats, with no significant differences between treatment with Ber or CCE and metformin or rosiglitazone. The indices of mean optical density and fasting ß­cell function index (FBCI) were different following treatment with Ber and CCE, compared with those in the model rats, which may have stimulated the pancreatic secretion of insulin. When Ber and CCE were used to examine the protective effects on Rin­5F cells, it was found that the Rin­5f cell GSIS, cell cycle, lipotoxic islet cell proliferation and protein expression of PARP­1 were altered and improved, which may have protected pancreatic islet ß­cells by improving islet ß­cell proliferation and the protein expression of PARP­1. CCE and Ber exerted similar effects when used for the treatment of T2MD rats, and may have stimulated the pancreatic secretion of insulin through the protective effect on islet ß­cells via improving islet ß­cell proliferation and the protein expression of PARP­1.

Poor permeability and absorption affect the activity of four alkaloids from Coptis.

Coptidis rhizoma (Coptis) and its alkaloids exert various pharmacological functions in cells and tissues; however, the oral absorption of these alkaloids requires further elucidation. The present study aimed to examine the mechanism underlying the poor absorption of alkaloids, including berberine (BER), coptisine (COP), palmatine (PAL) and jatrorrhizine (JAT). An ultra­performance liquid chromatography (UPLC) method was validated for the determination of BER, COP, PAL and JAT in the above experimental medium. In addition, the apparent oil­water partition coefficient (Po/w); apparent permeability coefficient (Papp), determined using a parallel artificial membrane permeability assay (PAMPA) plate; membrane retention coefficient (R %); and effect of P­glycoprotein (P­gp) inhibitor on the Papp of the four alkaloids were investigated. The intestinal absorption rate constant (Ka) and absorption percentage (A %) of the four alkaloids were also determined. The results of the present study demonstrated that the Po/w of the four alkaloids in 0.1 mol·l­1 HCl medium was significantly higher (P<0.01), compared with those of the alkaloids in phosphate buffer (pH 7.4). The Papp of BER was 1.0­1.2x10­6 cm·s­1, determined using a PAMPA plate, and the Papp of BER, COP, PAL and JAT decreased sequentially. The concentrations of the four alkaloids on the apical­to­basolateral (AP­BL) surface and the basolateral­to­apical (BL­AP) surface increased in a linear manner, with increasing concentrations between 10 and 100 µmol. In addition, the transportation of BER on the BL­AP surface was significantly faster (P<0.01), compared with that on the AP­BL surface and, following the addition of verpamil (a P­gp inhibitor), the Papp (AP­BL) of the four alkaloids increased, whereas the Papp (BL­AP) was significantly decreased (P<0.01). The rat intestinal perfusion experiment demonstrated that the four alkaloids were poorly absorbed; however, the Ka of BER was significantly higher, compared with the three other alkaloids. Furthermore, the A % and Ka provided evidence that the absorption of BER was increased in the jejunum, compared with in the ileum. In conclusion, the four alkaloids from Coptis appeared to be poorly absorbed, determined using a shake flask, pre­coated PAMPA plates, a Caco­2 cell monolayer model and intestinal perfusion; however, absorption was higher in the jejunum than in the ileum. Among the four alkaloids, the permeability of BER was markedly higher than the others, and P­gp efflux had a significant effect on the absorption of those alkaloids.

In vitro studies of berberine metabolism and its effect of enzyme induction on HepG2 cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Berberine (BER) and BER-original herbal medicines have a variety of pharmacological functions and have been widely used in clinical. However, its effect of enzyme induction on cytochrome P450 (CYP) in human hepatocytes is unknown. MATERIAL AND METHOD: Metabolism of berberine and its effect on main metabolic enzymes in HepG2 cell in vitro was investigated. Cocktail probe drugs, mRNA expression and protein expression were used to evaluate the metabolism potency. Meanwhile, an UPLC-MS/MS method was validated for the analysis of BER and four probe drugs in HepG2 cell. RESULT: BER significantly increased the metabolism of midazolam, phenacetin and tolbutamide by inducing the CYP1A2 and 3A4 enzyme in a dose-dependent manner, the mRNA and protein expression of CYP1A2 and 3A4 were increased by berberine at 1000ng·mL(-1). The activity of CYP1A2 and 3A4 could be induced by BER more than 500ng·mL(-1) in HepG2 cell, which was confirmed by the increase of its mRNA and protein expression. CONCLUSION: BER increases the metabolism of cocktail drugs such as midazolam, phenacetin and tolbutamide by increasing the mRNA and protein expression of CYP1A2 and 3A4.

Pharmacokinetics study of asperosaponin VI and its metabolites cauloside A, HN saponin F and hederagenin.

This experiment's with aim was to study the pharmacokinetics of asperosaponin VI and its three metabolites (cauloside A, HN saponin F and hederagenin) via a sensitive high performance liquid chromatography connected with electrospray ionization triple quadrupole mass spectrum (HPLC-ESI-MS/MS). Chromatographic separation was achieved on a reverse phase C18 column with a gradient mobile phase of CH3CN-water with 0.1 % HCOOH at a flow rate of 0.3 mL/min. Sample analysis was simultaneously performed with a multiple reaction monitoring mode using target determination ions at m/z 927.5 â†’ 603.4 for asperosaponin VI, m/z 811.1 â†’ 603.4 for cauloside A, m/z 649.4 â†’ 603.4 for HN saponin F, m/z 71.4 â†’ 393.3 for hederagenin and m/z 307.0 â†’ 161.1 for warfarin as the internal standard. The calibration curve was linear at the range of 0.25-500 ng/mL, and the lower limit of quantification was 0.25 ng/mL for each compound. While the precisely intra-assay and inter-assay variabilities were <9.5 and 7.8 %, respectively; accuracy was determined at the concentrations of 5, 25, 100 ng/mL for all the analytes with the relative standard deviation (%) no more than 15.0 %. Consequently, the validated method could be successfully and precisely applied to the pharmacokinetic study of asperosaponin VI and its metabolites. As a result, the pharmacokinetic parameters of cauloside A, HN saponin F and hederagenin such as T max were obtained at 9.33 ± 2.49, 7.33 ± 0.47 and 12.33 ± 2.36 h, respectively.

Validation of a HPLC-tandem MS/MS method for pharmacokinetics study of (+)-pinoresinol-di-ß-D-glucopyranoside from Eucommia ulmoides Oliv extract in rats' plasma.

Natural plant compounds have an unexceptional influence in pharmacy as they provide an uncountable number of invaluable lead molecules. Phytochemical researches nowadays focus on bio-assay guided revealing of the therapeutic profile and synergism of medicinal herbs and their constituents. Assessing the clinical and biological potential and determining the pharmacokinetics of herbal constituents is also an area of much interest. This work was conducted in order to carry out a sensitive liquid chromatography tandem mass spectrum (HPLC-MS/MS) method for the pharmacokinetics study of (+)-pinoresinol-di-ß-D-glucopyranoside (PG) in rats' plasma after oral administration of Eucommia ulmoides Oliv extract. The validated method was by means of linearity, precision, matrix effect and recovery so that it could be used for the pharmacokinetic study of PG. The obtained pharmacokinetic parameters shown that PG pertains to one-compartment model and 95% of PG was eliminated within 12h.

A peptide of foot-and-mouth disease virus serotype Asia1 generating a neutralizing antibody response, and an immunostimulatory peptide.

The epitopes of the capsid of foot-and-mouth disease virus (FMDV) play important roles in the construction of highly immunogenic subunit vaccines. However few epitopes have been found for FMDV serotype Asia1. In this study we screened for epitopes of the VP1 and VP2 proteins of FMDV serotype Asia1 isolate, YNBS/58. Fragments consisting of amino acids 133-163 of VP1 and amino acids 1-33 of VP2 contained epitopes, and both induced lymphoproliferation in guinea pigs. Only the VP1 fragment induced neutralizing antibodies but the VP2 peptide dramatically increased the neutralizing antibody response induced by the VP1 peptide.

[Cloning, expression of human keratinocyte growth factor and its purification and identification].

To clone KGF-2 gene, get hKGF-2 protein and detemine its activity. The cNDA of human KGF-2 was isolated from fetal lung by RT-PCR and cloned into pBV220 plasmid. The recombinant pBV220-hKGF-2 plasmid was transformed into E. coli (BL21), induced at 42 degrees C for the expression of hKGF-2. Recombinant human KGF-2 was purified from the ultrasonic-treated BL21 by heparin-Sepharose CL-6B treated column chromatography and cation exchange column chromatography. MTT method was used for the determination of its biological activity. SDS-PAGE showed that rhKGF-2 was expressed in E. coli BL21 as soluble protein of approximately 20kD. The rhKGF-2 protein can stimulate the proliferation of NIH3T3 cells significantly from 1 ng/mL to 10 ng/mL. HKGF-2 cDNA wasclned and highly expressed in E. coli BL21 and the purified rhKGF-2 showed the mitogenic activity on NIH3T3 cells.

Study on the mechanism of epidermal growth factor-induced proliferation of hepatoma cells.

AIM: Many growth factors, such as epidermal growth factor (EGF), are associated with the carcinogenesis. EGF plays its role in the proliferation of hepatoma cells through binding with EGF receptor (EGFR) and a series of signal transduction. But the postreceptor pathway is still not clear. In the present experiment, we studied the effect of tyrosine kinase, protein kinase C, Na(+)/H(+) exchange, calmodulin and voltage-dependent Ca(2+) channel on EGF-induced hepatoma cell proliferation. METHODS: Hepatoma cell line SMMC7721 was cultured in RPMI1640 serum-free medium. In order to study the effect of thyrosine kinase, protein kinase C, Na(+)/H(+) exchange, calmodulin and voltage-dependent Ca(2+) channel on human heptoma cell proliferation induced by epidermal growth factor (EGF), DNA synthesis rate of hepatoma cells was measured by the method of (3)H-TdR incorporation. RESULTS: EGF (10(-9) M) stimulated the proliferation of heptoma cells significantly ((3)H-TdR incorporation was 1 880+/-281 cpm/well, P<0.05), and this effect was significantly inhibited by tyrosine kinase inhibitor genistein ((3)H-TdR incorporation was 808+/-209 cpm/well, P<0.001). Calmodulin inhibitor W-7, protein kinase C inhibitor H-7 and Na(+)/H(+) exchange inhibitor amiloride individually had significant inhibiting effect on EGF-induced proliferation of hepatoma cells ((3)H-TdR incorporation was 978+/-87.3 cpm/well, 1 241+/-147 cpm/well, 1 380+/-189 cpm/well, respectively, P<0.001, P<0.01, P<0.05), but they all had no effect on the basal level proliferation of cultured hepatoma cells ((3)H-TdR incorporation was 1 284+/-260 cpm/well, 1 179+/-150 cpm/well, 1 392+/-152 cpm/well, respectively, (3)H-TdR incorporation of the control was 1 353+/-175 cpm/well, P>0.05). Voltage-dependent Ca(2+) channel inhibitor verapamil had no inhibition on EGF-induced proliferation of hepatoma cells ((3)H-TdR incorporation was 1 637+/-133 cpm/well, P>0.05), it also had no effect on the basal level proliferation of cultured hepatoma cells ((3)H-TdR incorporation was 1 196+/-112 cpm/well, P>0.05). CONCLUSION: Our data suggest that tyrosine kinase, Ca(2+)-calmodulin-dependent pathway, protein kinase C and Na(+)/H(+) exchange play a critical role in EGF-induced proliferation of hepatoma cells and that the effect of EGF is independent of voltage-dependent Ca(2+) channel.