Objective To screen B and T cell antigen epitopes on Acinetobacter baumannii outer membrane protein 33×103-36×103 (OMP33-36).Methods B and T cell epitopes on OMP33-36 of Acinetobacter baumannii were predicted by bioinformatics methods and synthesized.Recombinant expression plasmid pET-30a-OMP33-36 was cloned and used to express OMP33-36 in a prokaryotic expression system.The expressed OMP33-36 was used to immunize BALB/c mice after purification.Serum sample was collected from each mouse in immunization and negative control groups, and then analyzed by indirect ELISA with synthesized peptides to identify B cell epitopes.Splenocytes were separated from every mouse and then cultured with each of the synthesized peptides, respectively.Double sandwich ELISA was performed to detect IFN-γ secretion in the supernatant of cell cultures for screening of T cell epitopes.Results Candidates of B and T cell epitopes were constructed, which were PB1, PB2, PB3, PT1, PT2 and PT3.Results of the indirect ELISA showed that peptides PB1 and PB2 reacted with the serum samples collected from immunized mice and A450 values of the immunization group were significant higher than those of the negative control group.Compared with the negative control group, enhanced secretion of IFN-γ following peptide PT3 stimulation was observed in the immunization group as indicated by the double sandwich ELISA.Conclusion Two B cell epitopes PB1 and PB2, and one T cell epitope PT3 on the OMP33-36 of Acinetobacter baumannii were successfully constructed and screened out.

Background Blepharitis caused by Demodex infestation is very common in clinical practice.There are various methods mentioned in the study of Demodex infestation in China,but a unified introduction and evaluation of the operating procedures is lacked.A quick and accurate clinical diagnostic method for Demodex infestation needs to be further studied.Objective This study aimed to establish operation procedures for the clinical examination of eyelid Demodex infestation,which were applied to evaluate the conditions of eyelid Demodex infestation in ocular patients with discomfort.Methods One thousand and fifty-two patients with eye dryness,eye itchiness or other symptoms were selected for slit lamp examination and photographing of the eyelid margin area.Three eyelashes with associated scurf from each superior eyelid were plucked out for examination of Demodex under the microscope.Positive findings included observation of Demodex mites or eggs.Their amounts were recorded individually for all eyelash samples.Results A procedure for observing,recording and reporting eyelid Demodex infestations in patients was successfully established.By using this procedure,1 052 patients were investigated for the examination of Demodex infestations.Demodex mites or eggs were found in 582 cases (55.3％).The positive rate of Demodex infestation increased with age,and the population over 60 years group had the highest positive rate,showing a significant difference among the different age groups (x2=10.547,P=0.001).There was no significant difference in positive rate between male patients and female patients (P =0.352).The test turnaround time (TAT) for one examination was (11.4±5.2) seconds.Conclusions The operational procedure for examining the palpebral margin Demodex infestation by the slit-lamp,optical microscope,photographing and laboratory reports is established.It is simple and quick in the appliation for the clinical diagnosis of eyelid Demodex infestation.

Objective: To investigate whether Pseudomonas aeruginosa recombinant protein PA3611 can induce the in vitro epithelial-mesenchymal transformation (EMT) of rat bronchial epithelial cells. Methods: A series of processes including DNA synthesis, gene amplification, vector construction, induction of expression and protein purification was used to synthesize the recombinant protein PA3611. CCK-8 kit was used to detect the proliferation of bronchial epithelial cells after stimulation with the protein PA3611. Morphological changes in bronchial epithelial cells were observed. Western blot and qPCR were performed to measure the expression of E-cadherin (E-CAD) and α-smooth muscle actin (α-SMA). T test was used for statistical analysis. Results: DNA sequencing verified the successful preparation of the recombinant protein PA3611. PA3611 inhibited the proliferation (P<0.05) and induced the EMT of bronchial epithelial cells. Moreover, it also inhibited the expression of E-CAD protein, but promoted the expression of α-SMA (P<0.05). Conclusion The recombinant protein PA3611 can induce the EMT of bronchial epithelial cells and the underlying mechanisms are worthy of further investigation.

OBJECTIVETo evaluate the efficacy of periodontal treatment for the management of cardiovascular risk factors.

METHODSEligible studies in Cochrane Controlled Trials Register/CENTRAL, PubMed, EMBASE, and China Biology Medicine disc (CBMdisc) were searched until October 13, 2011. References of the included studies were hand searched. Two reviewers assessed the risk of bias and extracted the data of the included studies in duplicate. Meta-analysis was conducted with Revman 5.1.

RESULTSSix randomized controlled trials involving 682 participants were included. One case had low risk of bias, another one had moderate risk of bias, and the remaining four had high risk of bias. Meta-analysis showed that periodontal treatment has no significant effect on C-reactive protein, total cholesterol, low-density lipoprotein cholesterol, and triglycerides (P > 0.05). However, the treatment had a significant effect on high-density lipoprotein cholesterol [MD = 0.05, 95% CI (0.00, 0.09), P = 0.04].

CONCLUSIONPeriodontal treatment has good effects on controlling high-density lipoprotein cholesterol although more randomized controlled trials must be conducted to verify its effectiveness.

Objective:To explore the effect of macrophage-derived exosomes on the activation of hepatic stellate cells and its possible mechanism.Methods:Differential ultracentrifugation was used to extract macrophage exosomes. The exosomes were co-cultured with the mouse hepatic stellate cell line JS1, and a control group was established with phosphate buffered saline (PBS). Cell immunofluorescence was used to observe the expressional conditions of F-actin. Cell counting kit-8 (CCK8) was used to detect the survival rate of JS1 cells in the two groups. The activation indices of JS1 cells [collagen type Ⅰ (Col Ⅰ) and α-smooth muscle actin (α-SMA)] and its key signal pathway activation index expression level [transforming growth factor (TGF)-β1/Smads, platelet-derived growth factor (PDGF)] in the two groups were determined using Western blot and RT-PCR. Data comparison between two groups was performed using an independent sample t-test.Results:The membrane structure of exosomes was clearly observed by transmission electron microscopy. The expression of exosome marker proteins CD63 and CD81 was positive, suggesting that exosomes were successfully extracted. Exosomes were co-cultured with JS1 cells. Compared with the PBS control group, there was no statistically significant difference in the proliferation rate of JS1 cells in the exosomes group ( P＞0.05). The expression of F-actin was significantly increased in the exosome group. The mRNA and protein expression levels of α-SMA and ColⅠwere significantly increased in exosome group JS1 cells (all P＜0.05). The mRNA relative expression levels of α-SMA in PBS and exosome group were 0.25±0.07 and 1.43±0.19, respectively, while that of ColⅠ was 1.03±0.04 and 1.57±0.06, respectively. The mRNA and protein expressions of PDGF were significantly increased in exosome group JS1 cells ( P＜0.05). The mRNA relative expression levels of PDGF in the PBS group and exosome group were 0.27±0.04 and 1.65±0.12, respectively. There were no statistically significant differences in the mRNA and protein expressions of TGF-β1, Smad2 and Smad3 between the two groups ( P＞0.05). Conclusion:Macrophage-derived exosomes significantly promote the activation of hepatic stellate cells. JS1 cells may be the underlying mechanism for the up-regulation of PDGF expression.

This paper reports the progress of biodiesel production with enzymatic catalysis in Beijing University of Chemical Technology, one of the leaders in biodiesel R & D in China, which includes screening of high-yield lipase production strains, optimization and scale-up of the lipase fermentation process, lipase immobilization, bioreactor development and scale-up, biodiesel separation and purification and the by-product glycerol utilization. Firstly, lipase fermentation was carried out at industrial scale with the 5 m3 stirred tank bioreactor, and the enzyme activity as high as 8 000 IU/mL was achieved by the species Candida sp. 99-125. Then, the lipase was purified and immobilized on textile membranes. Furthermore, biodiesel production was performed in the 5 m3 stirred tank bioreactor with an enzyme dosage as low as 0.42%, and biodiesel that met the German biodiesel standard was produced. And in the meantime, the byproduct glycerol was used for the production of 1,3-propanediol to partly offset the production cost of biodiesel, and 76.1 g/L 1,3-propanediol was obtained in 30 L fermentor with the species Klebsiella pneumoniae.
Biofuels ; Bioreactors ; Biotechnology ; economics ; methods ; Candida ; enzymology ; Catalysis ; China ; Enzymes, Immobilized ; metabolism ; Esterification ; Fermentation ; Lipase ; metabolism ; Plant Oils ; chemistry

Objective To investigate the Rv2346c gene function through constructing Rv2346c gene knockout strains of Mycobacterium tuberculosis (M . tuberculosis) mediated by bacteriophage and observing its virulence after infecting mice lung tissue in vivo .Methods The affinal exchange sites (AES) of the target gene was built ,and then integrated into the phage genomes of M .tuberculosis for harvesting the phagemids .The phagemids was imparted into Mycobacterium smegmatis to get recombinant phages with the same AES .A high titer of the recombinant phages was harvested through amplification in vitro . The M .tuberculosis was transfected and coated on solid medium with hygromycin resistance and cultured for 4 weeks at 37℃ .Single clone was picked out and gene knock-out was confirmed by PCR . Then C57BL/6J mice were infected with either wild type strain (WT ) or knockout strain (KO ) of M . tuberculosis .Mice mortality ,lung tissue inflammation and colony-forming units (CFU ) counts in vitro were observed 6 to 8weeks post infection with different strains . Paired-samples t test was used for comparison between groups ,chi-square test was used for comparison of rates .Results The products of PCR and inserted fragment sizes were consisted with the expectation and confirmed to be the target gene . The target fragment of Rv2346c was removed successfully and the mice were infected for 6-8 weeks .Themice infected with Rv2346c KO strain had reduced mortality (53％ vs 20％ ,χ2 =6 .1112 ,P<0 .05) ,lung tissue inflammation (1040 ± 89 vs 1960 ± 56 ,t=7 .1016 ,P<0 .05) and CFU count in vitro (15 .0 ± 0 .8 vs 90 .0 ± 1 .5 ,t=23 .0361 , P<0 .05) compared with WT strain 6-8 weeks post infection .Conclusion Rv2346c gene knockout strains of M . tuberculosis mediated by bacteriophageis are successfully constructed ,which establishes the foundation for the future gene function study of Rv2346c .

Purpose To evaluate the function of swallowing by examining the relative motion of the hyoid bone and larynx(HL)via ultrasonography,and to explore the value of HL motion ratio in the evaluation of stroke with pharyngeal dysphagia.Materials and Methods A total of 43 stroke patients with dysphagia(dysphagia group)and 43 healthy adults(healthy control group)from June 2021 to April 2022 in the Third Affiliated Hospital of Henan University of Traditional Chinese Medicine and the First Affiliated Hospital of Henan University of Traditional Chinese Medicine were enrolled.The displacement and motion time of HL were measured by ultrasonography when the participants swallowed 5 ml water.The median flow tracking algorithm was implemented in Python language to measure the displacement of the hyoid bone,the movement time of the hyoid bone,the displacement of the larynx and the movement time of larynx,and then HL movement ratio was calculated,respectively.The differences in ultrasonography measurements between the two groups were compared,and the influencing factors of stroke with dysphagia were screened out via Logistic regression analysis.Then the receiver operating characteristic curve was drawn,the area under curve and the cut-off value,sensitivity and specificity were calculated subsequently.Results The larynx motion time(static phase),the larynx displacement(elevation phase)and the HL motion ratio were significantly related to swallowing in healthy participants,with significant differences between the two groups(t=4.97,6.38,6.17,P<0.05).The results of Logistic regression analysis showed that the HL motion ratio was the influencing factor of stroke with dysphagia(OR>1,P<0.05).The optimal cut-off value of the HL motion ratio for the diagnosis of dysphagia was 0.58,leading to a sensitivity of 86.0%,a specificity of 93.0%and the area under curve of 0.967,respectively.Conclusion Ultrasonography can quantitatively evaluate the motion of the HL during swallowing,and the HL motion ratio can be considered as a parameter for the evaluation of stroke with dysphagia,providing new insights for clinical diagnosis of dysphagia.

Nonalcoholic fatty liver disease(NAFLD)has developed into the most common chronic liver disease and can lead to liver cancer.Our laboratory previously developed a novel prescription for NAFLD,"Eight Zhes Decoction"(EZD),which has shown good curative effects in clinical practice.However,the pharmaco-dynamic material basis and mechanism have not yet been revealed.A strategy integrating lipidomics,network pharmacology and pharmacokinetics was used to reveal the active components and mecha-nisms of EZD against NAFLD.The histopathological results showed that EZD attenuated the degrees of collagen deposition and steatosis in the livers of nonalcoholic steatofibrosis model mice.Furthermore,glycerophospholipid metabolism,arachidonic acid metabolism,glycerolipid metabolism and linoleic acid metabolism with phospholipase A2 group IVA(PLA2G4A)and cytochrome P450 as the core targets and 12,13-cis-epoxyoctadecenoic acid,12(S)-hydroxyeicosatetraenoic acid,leukotriene B4,prostaglandin E2,phosphatidylcholines(PCs)and triacylglycerols(TGs)as the main lipids were found to be involved in the treatment of NAFLD by EZD.Importantly,naringenin,artemetin,canadine,and bicuculline were iden-tified as the active ingredients of EZD against NAFLD;in particular,naringenin reduces PC consumption by inhibiting the expression of PLA2G4A and thus promotes sufficient synthesis of very-low-density lipoprotein to transport excess TGs in the liver.This research provides valuable data and theoretical support for the application of EZD against NAFLD.

Xiaoji Yinzi is one of the classic prescriptions for treating urinary diseases， originated from the Yan's Prescriptions to Aid the Living （Yan Shi Ji Sheng Fang） written by YAN Yonghe in the Song dynasty. Xiaoji Yinzi is composed of Rehmanniae Radix， Cirsii Herba， Talcum， Akebiae Caulis， Typhae Pollen， Nelumbinis Rhizomatis Nodus， Lophatheri Herba， Angelicae Sinensis Radix， Gardeniae Fructus， and Glycyrrhizae Radix et Rhizoma and has the effects of cooling blood and stopping bleeding， draining water and relieving stranguria. The medical experts of later generations have inherited the original prescription recorded in the Yan's Prescriptions to Aid the Living， while dispute has emerged during the inheritance of this prescription. In this study， the method of bibliometrics was employed to review and analyze the ancient documents and modern clinical studies involving Xiaoji Yinzi. The results showed that Xiaoji Yinzi has two dosage forms： powder and decoction. According to the measurement system in the Song Dynasty， the modern doses of hers in Xiaoji Yinzi were transformed. In the prepration of Xiaoji Yinzi powder， 149.2 g of Rehmanniae Radix and 20.65 g each of Cirsii Herba， Talcum， Akebiae Caulis， stir-fried Typhae Pollen， Nelumbinis Rhizomatis Nodus， Lophatheri Herba， wine-processed Angelicae Sinensis Radix， stir-fried Gardeniae Fructus， and stir-fried Glycyrrhizae Radix et Rhizoma are grounded into fine powder with the particle size of 4-10 meshes and a decocted with 450 mL water to reach a volume of 240 mL. After removal of the residue， the decoction was taken warm before meals， 3 times a day （i.e.， 7.77 g Rehmanniae Radix and 0.97 g each of the other herbs each time）. In the preparation of Xiaoji Yinzi decoction， 20.65 g each of the above 10 herbs are used， with stir-fried Typhae Pollen， wine-processed Angelica Sinensis Radix， stir-fired Gardeniae Fructus， stir-fired Glycyrrhizae Radix et Rhizoma， and raw materials of other herbs. Xiaoji Yinzi is specialized in treating hematuresis and blood stranguria due to heat accumulation in lower energizer， which causes injury of the blood collaterals of gallbladder and dysfunction of Qi transformation. In modern clinical practice， Xiaoji Yinzi is specifically used for treating urinary diseases and can be expanded to treat diseases of the cardiovascular system and other systems according to pathogenesis. The comprehensive research on the key information could provide a scientific reference for the future development of Xiaoji Yinzi.