DNA binding properties and cell viabilities of metal complexes derived from (E)-2-hydroxy-N'-((thiophen-2-yl)methylene)benzohydrazone and (E)-N'-((thiophen-2-yl)methylene)isonicotinylhydrazone.

OBJECTIVE: This study aims to investigate the CT-DNA (Calf thymus DNA) binding properties and HeLa cell viabilities of metal complexes derived from (E)-2-hydroxy-N'-((thiophen-2-yl)methylene)benzohydrazone (H2L1) and (E)-N'-((thiophen-2-yl)methylene)isonicotinylhydrazone (HL2). MATERIALS AND METHODS: A series of metal complexes derived from (E)-2-hydroxy-N'-((thiophen-2-yl)methylene)benzohydrazone (H2L1) and (E)-N'-((thiophen-2-yl)methylene)isonicotinylhydrazonewere (HL2) were synthesized, and their structures were characterized through FT-IR, ESI-MS, elemental analysis, molar conductivities and X-ray diffraction. DNA binding properties between CT-DNA and metal complexes were investigated by UV-Vis spectrophotometry and viscosity titration. The toxicological properties of compounds on HeLa cell were measured in vitro. RESULTS: Ligand H2L1 or HL2 exhibits a tridentate and anion ligand and uses oxygen anion, nitrogen atom and sulfur atom to coordinate with metal ions. When coordinated with metal ions, the unit O=C-NH- of each ligand has been enolized and deprotonated into -O-C=N-. The suggested chemical formulas of metal complexes are: [Co(HL1)2], [Ni(HL1)2], [Cu(HL1)2], [Co(L2)2], [Cu(L2)2], [Zn(L2)2], [ScL2(NO3)2(H2O)2], [Pr(L2)2(NO3)] and [Dy(L2)2(NO3)]. Both ligands and their metal complexes can bind strongly to CT-DNA through hydrogen bond and intercalation with Kb of 104~105 L mol-1 compared to ethidium bromide [classical DNA intercalator, Kb(EB-DNA) = 3.068 × 104 L mol-1]; however, the groove pattern cannot be excluded. The coexistence of multiple binding modes may be a common form of drug binding to DNA. HeLa cell shows lower viabilities in the presence of [Ni(HL1)2] and [Cu(HL1)2] (*p < 0.05) compared to the other compounds, with the LC50 of 2.6 µmol L-1 and 2.2 µmol L-1, respectively. CONCLUSIONS: These compounds, especially [Ni(HL1)2] and [Cu(HL1)2], will be promising for anti-tumor drugs, which should be further studied.

[Predictive value of ultrasound-guided anesthesia injection in arthroscopy for borderline developmental dysplasia of the hip].

Objective: To evaluate the value of ultrasound-guided intra-articular anesthetic injection in predicting postoperative outcomes for borderline developmental hip dysplasia (BDDH). Methods: A follow-up study. The clinical data of 37 BDDH patients who received ultrasound-guided intra-articular anesthetic injection and arthroscopic examination in the Department of Sports Medicine, Senior Department of Orthopedics, the Fourth Medical Center of PLA General Hospital from May 2018 to February 2021 were retrospectively analyzed. Among them, there were 17 males and 20 females with a mean age of (37.9±12.8) years. All patients underwent ultrasound-guided intra-articular anesthetic injection prior to arthroscopy, and were evaluated with hip physical examination before and after injection, as well as before and after arthroscopy, in order to obtain the visual analog score (VAS) of pain for seven assessments. The total VAS score was calculated based on these evaluations. Follow-up was conducted for at least 12 months. The effective rate of injection referred to the ratio of the improvement of VAS score after anesthetic injection to the total VAS score before injection. Pearson correlation analysis and Bland-Altman analysis were used to test the correlation between modified Harris hip score (mHHS) after ultrasound-guided intra-articular anesthetic injection and mHHS score after arthroscopic surgery. A binary logistic regression model was established to analyze the substantial clinical benefit (SCB) for patients. Following the logistic regression analysis, a receiver operating characteristic (ROC) curve was constructed to evaluate the predictive power of ultrasound-guided intra-articular anesthetic injection in achieving SCB in those patients. The optimal cut-off value for injection efficacy was determined based on the ROC curve when SCB was achieved. Results: The follow-up time for all patients was (26.3±7.6) months. After anesthetic injection for 20 minutes, the total VAS score of pain [M(Q1,Q3)] decreased from 13(8,23) points before injection to 1(0,4) points; and the mHHS score [M(Q1,Q3)] increased from 60(46,70) points before arthroscopy to 90(84,96) points after, with statistically significant differences before and after injection and before and after arthroscopy (both P<0.001). Pearson correlation analysis showed that the mHHS score after intra-articular anesthetic injection was positively correlated with the mHHS score after surgery (r=0.961, P<0.001). The area under the ROC curve for predicting SCB after arthroscopy with ultrasound-guided intra-articular anesthetic injection was 0.769 (95%CI: 0.561-0.976), the Youden index was 0.663, the cut-off value was 0.569 2, the sensitivity was 96.3%, and the specificity was 70.0%. Conclusions: The results of ultrasound-guided intra-articular anesthetic injection before arthroscopy can indicate the presence of intra-articular lesions, and the degree of pain relief after injection is proportional to the functional recovery after arthroscopy. Patients with intra-articular anesthetic injection efficacy>56.92% have better results in hip arthroscopy.

[Clinical efficacy of the treatment of bilateral gluteal muscle contracture by inside-out iliotibial band release under arthroscopy in the supine position].

Objective: To investigate the clinical efficacy of bilateral gluteal muscle contracture treated with inside-out iliotibial band release under arthroscopy in the supine position. Methods: A prospective non-randomized controlled trial. Forty-six patients admitted to the Department of Sports Medicine, Senior Department of Orthopedics, the Fourth Medical Center of PLA General Hospital from April 2021 to August 2022 for bilateral gluteal muscle contracture and proposed surgical treatment were enrolled. The subjects were divided into two groups according to the preferred surgical protocols of the patients: the supine position group was treated with inside-out iliotibial band release under arthroscopy in the supine position, and the operation in lateral position group was carried out with outside-in iliotibial band release under arthroscopy in the lateral position. The total duration of non-surgical operations and the total duration of surgical operations were recorded for all patients. The gluteal muscle contracture disability scale within 3 days before surgery and at least 2 months after surgery were compared between the two groups, and the occurrence of complications between the two groups was compared too. Results: There were 26 cases in the supine position group, 11 males and 15 females with a mean age of (31.8±7.3) years; and there were 20 cases in the lateral position group, 7 males and 13 females with a mean age of (30.6±6.3) years. The differences in gender, age, body mass index (BMI) and postoperative follow-up time between the two groups were not statistically significant (all P>0.05). The total duration of non-surgical operations was shorter in the supine position group than in the lateral position group [(47.9±10.4) min vs (63.9±7.5) min, P<0.001]. There was no statistically significant difference in the total duration of surgical operations between the supine position group and the lateral position group [31.0(27.0, 43.5) min vs 33.0(24.8, 38.0) min, P>0.05]. The postoperative gluteal muscle contracture disability scales were significantly improved in both the supine position and lateral position groups when compared with those before the operation [93.0 (85.0, 98.0) vs 61.0 (50.5, 66.8), P<0.001 and 88.5±6.9 vs 63.6±9.6, P<0.001, respectively]. There was no statistically significant difference in the gluteal muscle contracture disability scale between the supine position and lateral position groups before and 2 months after surgery [59.3±11.9 vs 63.6±9.6 and 93.0 (85.0, 98.0) vs 89.5(84.0, 94.8), both P>0.05, respectively]. Two patients in each group developed subcutaneous hematoma after surgery, and all of them resolved within 2 weeks after surgery, the difference in complication incidence rate was not statistically significant (P>0.05). No postoperative complications such as fat liquefaction in the operated area, infection, decreased hip abductor muscle strength or nerve injury in the lower extremity were observed in both groups. Conclusion: The treatment of bilateral gluteal muscle contracture by inside-out iliotibial band release under arthroscopy in the supine position can effectively improve clinical efficiency, with definite efficacy, and it is an operative program worth promoting.

[Schistosomiasis control effect in Mianyang City in ten years after Wenchuan earthquake].

OBJECTIVE: To evaluate the effect of schistosomiasis control in Mianyang City in ten years after Wenchuan earth-quake, so as to provide the experiences for schistosomiasis control post-disaster. METHODS: The data of implementation of schistosomiasis control work in ten years after the Wenchuan earthquake were collected and analyzed for the epidemic situation of schistosomiasis in Mianyang City. Meanwhile, the awareness situation on schistosomiasis control of villagers and students was investigated by questionnaires in 2008 and 2015. RESULTS: All of the 6 counties (cities, districts) with schistosomiasis endemic in Mianyang City were hit by the earthquake disaster. After the disaster, the measurements including the conventional schistosomiasis control measures, the control of exogenous infection sources, the control of Oncomelania hupensis snails, and health education were carried out. The relevant departments of schistosomiasis control were cooperated to implement the prevention and control measures. The schistosomiasis prevalence of population and the snail condition rose in the year of the earthquake, but then declined year by year. In 2015, the awareness rates of schistosomiasis control of both villagers and the students were significantly improved compared with those in 2008. CONCLUSIONS: The measures of schistosomiasis control after the earthquake disaster are effective in Mianyang City, and the goal to prevent major plague after the earthquake is achieved.

Genome-wide differentially methylated genes in prostate cancer tissues from African-American and Caucasian men.

Increasing evidence suggests that aberrant DNA methylation changes may contribute to prostate cancer (PCa) ethnic disparity. To comprehensively identify DNA methylation alterations in PCa disparity, we used the Illumina 450K methylation platform to interrogate the methylation status of 485,577 CpG sites focusing on gene-associated regions of the human genome. Genomic DNA from African-American (AA; 7 normal and 3 cancers) and Caucasian (Cau; 8 normal and 3 cancers) was used in the analysis. Hierarchical clustering analysis identified probe-sets unique to AA and Cau samples, as well as common to both. We selected 25 promoter-associated novel CpG sites most differentially methylated by race (fold change > 1.5-fold; adjusted P < 0.05) and compared the ß-value of these sites provided by the Illumina, Inc. array with quantitative methylation obtained by pyrosequencing in 7 prostate cell lines. We found very good concordance of the methylation levels between ß-value and pyrosequencing. Gene expression analysis using qRT-PCR in a subset of 8 genes after treatment with 5-aza-2'-deoxycytidine and/or trichostatin showed up-regulation of gene expression in PCa cells. Quantitative analysis of 4 genes, SNRPN, SHANK2, MST1R, and ABCG5, in matched normal and PCa tissues derived from AA and Cau PCa patients demonstrated differential promoter methylation and concomitant differences in mRNA expression in prostate tissues from AA vs. Cau. Regression analysis in normal and PCa tissues as a function of race showed significantly higher methylation prevalence for SNRPN (P = 0.012), MST1R (P = 0.038), and ABCG5 (P < 0.0002) for AA vs. Cau samples. We selected the ABCG5 and SNRPN genes and verified their biological functions by Western blot analysis and siRNA gene knockout effects on cell proliferation and invasion in 4 PCa cell lines (2 AA and 2 Cau patients-derived lines). Knockdown of either ABCG5 or SNRPN resulted in a significant decrease in both invasion and proliferation in Cau PCa cell lines but we did not observe these remarkable loss-of-function effects in AA PCa cell lines. Our study demonstrates how differential genome-wide DNA methylation levels influence gene expression and biological functions in AA and Cau PCa.

Characterization of a Saccharomyces cerevisiae mutant with oversecretion phenotype.

An oversecreting mutant of Saccharomyces cerevisiae was obtained from about 400 meiotic segregants derived from thediploid cells made by crossing the HBsAg-induced mutant NI-C with the wild-type strain Sey6211. When transformed with a plasmid containing mouse alpha-amylase cDNA, the mutant (NI-C-D4) exhibited an increased capacity (up to 13-fold) for the secretion of mouse alpha-amylase, higher than the parental strains and other standard wild-type strains. It was also shown that alpha-amylase secreted by the oversecreting mutant had a higher activity and contained more of the non-glycosylated form than the glycosylated form. This isolated oversecreting, low-glycosylation mutant may prove to be a potential S. cerevisiae host for the production of foreign proteins. Further genetic analysis suggested that the mutation responsible for the mutant's oversecretion was partially dominant and that both the oversecretion and low-glycosylation phenotypes were governed by a single chromosome mutation. These pleiotrophic phenotypes may be attributed to a defect in the synthesis of an ER-resident chaperone.

Induction of a mitosis delay and cell lysis by high-level secretion of mouse alpha-amylase from Saccharomyces cerevisiae.

Some foreign proteins are produced in yeast in a cell cycle-dependent manner, but the cause of the cell cycle dependency is unknown. In this study, we found that Saccharomyces cerevisiae cells secreting high levels of mouse alpha-amylase have elongated buds and are delayed in cell cycle completion in mitosis. The delayed cell mitosis suggests that critical events during exit from mitosis might be disturbed. We found that the activities of PP2A (protein phosphatase 2A) and MPF (maturation-promoting factor) were reduced in alpha-amylase-oversecreting cells and that these cells showed a reduced level of assembly checkpoint protein Cdc55, compared to the accumulation in wild-type cells. MPF inactivation is due to inhibitory phosphorylation on Cdc28, as a cdc28 mutant which lacks an inhibitory phosphorylation site on Cdc28 prevents MPF inactivation and prevents the defective bud morphology induced by overproduction of alpha-amylase. Our data also suggest that high levels of alpha-amylase may downregulate PPH22, leading to cell lysis. In conclusion, overproduction of heterologous alpha-amylase in S. cerevisiae results in a negative regulation of PP2A, which causes mitotic delay and leads to cell lysis.

Asparagine as a nitrogen source for improving the secretion of mouse alpha-amylase in Saccharomyces cerevisiae protease A-deficient strains.

A modified chemically defined medium was achieved by using asparagine as a nitrogen source to increase the production of secreted mouse alpha-amylase in several Saccharomyces cerevisiae protease A-deficient (pep4) strains. The specific productivity (quantity) and the 53 kDa non-glycosylated active form (quality) of mouse salivary alpha-amylase in liquid medium containing asparagine was remarkably improved compared to media containing other nitrogen sources, including ammonium sulphate, glutamic acid, arginine, casamino acids, yeast extract and peptone. Similar improvement was also observed on starch solid agar regarding the clarity and size of the halo zone formed by alpha-amylase activity. Compared with ammonium sulphate, advantages of using asparagine as the nitrogen source in liquid or solid medium included increasing the cell mass of test strains, recovering the viability of protease-deficient strains to levels similar to the wild-type strain, and increasing the copy number of the mouse alpha-amylase expression vector in test strains. In turn, these advantages apparently contributed to the increase of secretion of mouse alpha-amylase in several test strains and especially in the protease A-deficient strains. In addition to demonstrating the use of modified chemically defined medium to improve the quality and quantity of secreted mouse alpha-amylase, this study also provides a new strategy to improve the secretion of heterologous proteins in protease A deficient strains.