Objective: To clone the full-length cDNA encoding squalene epoxidase (SE), which is the key enzyme involved in the triterpenoid biosynthesis pathway in Inonotus baumii, and analyze its bioinformates. Methods: Taking total RNA as template, the full length cDNA and DNA of SE in I. baumii was cloned through RT-PCR and the rapid amplification of cDNA ends (RACE) technique. The bioinformatics of SE gene were analyzed by ExPASy on line. Results: Sequence analysis showed that it consisted of 2145 bp with an open reading frame (ORF) of 1 452 bp, encoding 483 amino acid polypeptides. SE gene contained six exons and five introns. The relative molecular mass of SE calculated was 5.3 × 104, the isoelectric point (pI) was 8.41, and there was no signal peptide in SE. Conclusion: It is the first report that the cDNA encoding SE from I. baumii is cloned. This work provides a scientific basis for exploring the triterpenoid biosynthesis pathway of the medicinal ingredient and improving its quality in I. baumii.

The quality control of Chinese materia medica (CMM) has been the focus of CMM modernization, and how to demonstrate the consistency of product quality and clinical efficacy has become an important research aspect of development and innovation of CMM. Due to the heterogeneity of CMM, it is necessary to combine biopotency with other detection methods to guarantee the quality of medicines. This article reviewed the research status of CMM biopotency, analyzed the global local government regulations about biopotency and the difficulties of development in CMM, and discussed development ideas of biopotency based on PK-PD for CMM. Its main aim is to provide reference for the construction of the follow-up quality evaluation system, and to promote recognition from the international drug administration management on CMM quality standard.

Very low-density lipoprotein receptor(VLDLR)is a transmembrane lipoprotein receptor and plays an important role in the disorder of lipid metabolism in diabetes.The finding of changes in expression and distribution of two subtypes of VLDLR in diabetes as well as the effective VLDLR gene therapy in hyperlipidemia have drawn attention to the VLDLR as a potential target for prevention of abnormal lipid metabolism in diabetes.

Mycoplasma pneumoniae infection can cause coagulation dysfunction and thrombosis, involving organ including deep vein, lung and brain, etc.Central venous catheterization is the most important risk factor for deep venous thrombosis.Lung consolidation more than 2/3 is a high risk of pulmonary embolismin mycoplasma pneumoniae pneumonia.Elevated blood D-dimer over 5 mg/L is an independent risk factor for predicting thrombus.Color Doppler ultrasound and CT angiography(CTA) are the main methods to determine thrombosis.Deep venous thrombosis or pulmonary embolism should be treated with individualized anticoagulant and thrombolytic therapy.

OBJECTIVE:To explore the method of pharmaceutical care for antifungal drug treatment of new type crypto-coccal meningitis by clinical pharmacist. METHODS:Clinical pharmacist participated in the drug treatment process for a pa-tient with new type cryptococcal meningitis. Clinical pharmacist provided pharmaceutical care in following aspects:assisting doctor to optimize antifungal drugs treatment plan,providing patients pharmaceutical monitoring and medication education, etc. During amphotericin B treatment,the patient developed refractory hypokalemia. Clinical pharmacists suggested doctors to reduce the dose of amphotericin B and additionally use voriconazole for antifungal treatment. RESULTS:The patient devel-oped refractory hypokalemia no more after the plan was adjusted. After 11 weeks of systematic antifungal treatment,the pa-tient was on the mend. CONCLUSIONS:The participation of clinical pharmacist in antifungal treatment of new type cryptococ-cal meningitis indicates that following the instructions,but not lost flexible disposal;providing service actively,and details is guarantee of safety.

OBJECTIVETo study cellular and humoral immune status on prophase of severe hepatitis B (PSHB).

METHODS56 cases of PSHB patients, 40 cases of chronic hepatitis B (CHB) patients and 20 cases of healthy volunteers were enrolled for detection of CD3+, CD4+, CD8+ and CD3-/CD19+ (B cells) lymphocyte subsets in peripheral blood by flow cytometry. Serum IgG and complement C3 was detected by immunoturbidimetry and analyzed statistically.

RESULTSCompared with CHB group and healthy control group, percentage of lymphocyte subsets CD8+ were significantly lower in PSHB group (P < 0.01 or P < 0.05). While the percentage of lymphocyte subsets CD4+ and ratio of CD4+/CD8+ in PSHB group was obviously higher than those in CHB group (P < 0.+01 or P < 0.05). In addition, There was no significant difference on the percentage of B cell and level of serum IgG between PSHB group and CHB group (P > 0.05, while the level of serum complement C3 in PSHB group were significantly lower than those in CHB group and healthy control group (P < 0.01, P < 0.05).

CONCLUSIONPSHB has a certain degree of cellular immune dysfunction, which characterized by cellular immune function hyperfunction and humoral immune suppression.

Adult ; Antibodies, Viral ; immunology ; Complement C3 ; immunology ; Female ; Flow Cytometry ; Hepatitis B, Chronic ; immunology ; Humans ; Lymphocyte Count ; Male ; Middle Aged ; T-Lymphocyte Subsets ; cytology ; immunology ; Young Adult

Objective: To clone and characterize a 3-hydroxy-3-methylglutaryl-CoA synthase (HMGS) gene which involved in the triterpenoid biosynthesis pathway in Sanghuangporus baumii. Methods: The HMGS gene cDNA full-length sequence was cloned by RACE technology. Characteristics including the physicochemical properties and conserved domain of the deduced HMGS protein were determined by a series of bioinformatics tools. The entire protein-coding cDNA of HMGS was cloned into the expression vector pET-32a (+). Then the recombinant plasmid was transformed into E. coli BL21 (DE3) cells. With IPTG induction, SDS-PAGE was used to investigate the situation of expression. Additionally, qRT-PCR technology was performed to measure the transcript levels of HMGS gene in the triterpenoid pathway during different developmental stages of S. baumii. Results: The full-length nucleotide sequence of HMGS was 1 930 bp, containing a complete open reading frame of 1 458 bp which encoded a polypeptide of 485 amino acids. Bioinformatics analysis of the amino acid sequence showed that the molecular weight of encoded protein was 52 750, and theoretical isoelectric point was 5.60. This protein was a hydrophilic protein, without transmembrane and signal peptide sequence. The constructed phylogenetic tree showed that HMGS from S. baumii had the highest similarity with HMGS from Fomitiporia mediterranea. The prokaryotic expression vector pET-32a-HMGS was sucessfully obtained. SDS-PAGE results showed that a significant protein band was in consistent with molecular weight of the predicted protein. Moreover, the results showed that the transcript levels of HMGS gene were in dynamic change. The transcript levels in the mycelium stage were higher than that in the fruiting body stage. For instance, the highest transcript level of HMGS was at 14 d and was 2.33-fold higher than the 5 d. Conclusion: Molecular characterization of HMGS will be useful for further functional elucidation of the gene involving in triterpenoid biosynthesis pathway in S. baumii.

Objective To obtain prokaryotic expression and over-expression vectors of squalene epoxidase (SE) gene from Sanghuangporus baumii. Methods The entire protein-coding cDNA of SE was cloned into the expression vector pET-32a (+). Then the recombinant plasmid was transformed into Escherichia coli BL21 (DE3) cells. SDS-PAGE was used to investigate the situation of expression after IPTG induction for 2-10 h. Additionally, primers were designed according to the gpd promoter sequence of Lentinula edodes in GenBank, and the gpd promoter fragment was obtained by PCR. Subsequently, the plant binary expression vector pCAMBIA1301 was selected as the basic vector, and then the 35 S promoter replaced with L. edodes gpd promoter through enzyme digestion and connection. Finally, the coding region of SE was cloned to the downstream of the gpd promoter to construct over-expression vectors. Results The prokaryotic expression vector pET-32a-SE was successfully obtained. SDS-PAGE results showed a significant protein band was found in the vicinity of the relative molecular weight of approximately 55 000, consistent with molecular weight of the predicted protein. Moreover, the over-expression vector pCAMBIA1301-gpd-gpd-SE was constructed successfully through different detection ways. Conclusion These results lay the foundation for the further study of SE in triterpenoid biosynthesis pathway of S. baumii.

Background The pathogenesis of pterygium has been in the study.Relative molecular biology study showed that pterygium is a tumor-like lesion,and based on overseas literatures,human papillomavirus (HPV) is positively expressed in 100％ patients with pterygium in some region.However,if this result is suitable for Chinese patients is unclear.Objective This study was to identify the role of human HPV in the pathogenesis of pterygia in Wuxi area.Methods Forty-eight pterygium specimens including 7 recurrent pterygia and 41 primary pterygia were collected during the operation,and these patients were from Wuxi area.Two cervical carcinoma specimens and 2 conjunctiva specimens from normal donors were obtained as positive control and negative control respectively.The fluorescent quantitative polymerase chain reaction (FQ-PCR) was used to detect HPV DNA of specimens.Results The amplified curves of HPV 6,11 of pterygium specimens,cervical carcinoma specimens and normal specimens were all below the positive quality control curve,but the amplified curves of HPV16,18 were above the quality control curve in cervical carcinoma specimens; while those of pterygium specimens and normal conjunctival specimens were all below the quality control curve.HPV16/18 was identified in 2 cervical carcinoma specimens,but no HPV6/11 was detected in 2 cervical carcinoma specimens.However,HPV DNA expression in primary and recurrent pterygias were absent.Conclusions According to these results,HPV is not the primary cause for the pathogenesis of pterygium in Wuxi region.

AlM:To compare the functional and cosmetic effects of two different surgical techniques for congenital ptosis.
METHODS: The patients were divided into four groups according to the operation method: Patients undertook bilateral fascial suspension surgery as Group A ( 42 eyes of 21 cases ); Patients undertook bilateral levator muscle shortening surgery as Group B ( 38 eyes of 19 cases );Patients undertook unilateral fascial suspension surgery as Group C ( 24 eyes of 24 cases ); Patients undertook unilateral levator muscle shortening surgery as Group D (29 eyes of 29 cases). Each group patients were followed for postoperative function and appearance effect.
RESULTS: 1 ) Early postoperative of two operation function success rate was up to 100%, the function of levator muscle shortening surgery was 97. 01% in the late postoperative, was higher than bilateral fascial suspension surgery (87. 88%), with statistical difference in both surgerys (P<0. 05). 2) Appearance effect of two kinds of operation method in early postoperative had no statistical difference (P >0. 05); ln the late postoperative, the mean grades for “Lid Contour” and “Lid Crease” of Group B were better than that of Group A (P<0. 01). While the mean grades for “Lid Contour”, “Symmetry of Lid Height” and “Lid Crease” of Group D were similarly better than that of Group C (P<0. 01).
CONCLUSlON: Two kinds of operation method have good effects on the treatment of congenital ptosis. ln terms of cosmetic effect, levator muscle shortening surgery is better.