The ubiquitin ligase human TRIM71 regulates let-7 microRNA biogenesis via modulation of Lin28B protein.

let-7 microRNA (miRNA) is implicated in various biological processes, and its downregulation essentially linked to human malignancy. Regulation of gene expression of the let-7 family is critically linked to RNA-binding proteins. For instance, Lin28B and its paralog, Lin28A, inhibit the pre-let-7 precursor from being processed to mature miRNA by recruiting terminal uridyltransferase, TUT4, which adds oligomeric U at the 3' end, suggesting that deregulation of Lin28B, together with Lin28A, may alter various biological processes through modulation of let-7 expression. Here, we showed that the Lin28B protein level is regulated via ubiquitin-mediated proteasomal degradation, and identified the ubiquitin ligase as human TRIM-NHL domain-containing TRIM71. In cells, TRIM71 negatively regulates Lin28B protein stability by catalyzing polyubiquitination. Compared with its paralog, Lin28A, a C-terminal unique ~50 amino acid stretch of Lin28B is essential for TRIM71 interactions and subsequent polyubiquitination. Moreover, the N-terminal RING finger motif of TRIM71 is critical for protein-protein interactions and polyubiquitination of Lin28B, and consequent let-7 expression. Consistent with the let-7 stimulatory role of TRIM71 via Lin28B polyubiquitination, specific knockdown of TRIM71 led to downregulation of let-7 expression. Expression of one of the known let-7 targets, HMGA2, was derepressed after knockdown of TRIM71. We additionally showed that enhanced expression of let-7 is part of a feedback loop that targets TRIM71 3'UTR, which contains two conserved let-7 target sites. Our findings collectively reveal critical aspects of regulatory complexity of let-7 biogenesis at the posttranscriptional level.

Expression of phosphoenolpyruvate carboxykinase linked to chemoradiation susceptibility of human colon cancer cells.

BACKGROUND: Resistance to 5-fluorouracil (5-FU) in patients with colorectal cancer prevents effective treatment and leads to unnecessary and burdensome chemotherapy. Therefore, prediction of 5-FU resistance is imperative. METHODS: To identify the proteins linked to 5-FU resistance, two-dimensional gel electrophoresis-based proteomics was performed using the human colon cancer cell line SNU-C4R with induced 5-FU resistance. Proteins showing altered expression in SNU-C4R were identified by matrix-associated laser desorption/ionization-time-of-flight analysis, and their roles in susceptibility to 5-FU or radiation were evaluated in various cell lines by transfection of specific siRNA or creation of overexpression constructs. Changes in cellular signaling and expression of mitochondrial apoptotic factors were investigated by Western Blot analysis. A mitochondrial membrane potential probe (JC-1 dye) and a flow cytometry system were employed to determine the mitochondrial membrane potential. Finally, protein levels were determined by Western Blot analysis in tissues from 122 patients with rectal cancer to clarify whether each identified protein is a useful predictor of a chemoradiation response. RESULTS: We identified mitochondrial phosphoenolpyruvate carboxykinase (mPEPCK) as a candidate predictor of 5-FU resistance. PEPCK was downregulated in SNU-C4R compared with its parent cell line SNU-C4. Overexpression of mPEPCK did not significantly alter the susceptibility to either 5-FU or radiation. Suppression of mPEPCK led to a decrease in both the cellular level of phosphoenolpyruvate and the susceptibility to 5-FU and radiation. Furthermore, the cellular levels of phosphoenolpyruvate (an end product of PEPCK and a substrate of pyruvate kinase), phosphorylated AKT, and phosphorylated 4EBP1 were decreased significantly secondary to the mPEPCK suppression in SNU-C4. However, mPEPCK siRNA transfection induced changes in neither the mitochondrial membrane potential nor the expression levels of mitochondrial apoptotic factors such as Bax, Bcl-2, and Bad. Downregulation of total PEPCK was observed in tissues from patients with rectal cancer who displayed poor responses to preoperative 5-FU-based radiation therapy. CONCLUSION: Our overall results demonstrate that mPEPCK is a useful predictor of a response to chemoradiotherapy in patients with rectal cancer.

SRF-rearranged cellular perivascular myoid tumor: a clinicopathological analysis of two cases / 中华病理学杂志

Objective: To investigate the clinicopathological features, immunophenotype, diagnosis and differential diagnosis of SRF-rearranged cellular perivascular myoid tumor. Methods: Two cases of SRF-rearranged cellular perivascular myoid tumor diagnosed in the Department of Pathology, Fudan University Shanghai Cancer Center from October 2021 to March 2022 were collected. Immunohistochemical staining, fluorescence in-situ hybridization (FISH) and next-generation sequencing (NGS) were performed, and the literature was reviewed. Results: Case 1, a 3-month-old boy presented with a painless tumor of the scalp, measuring about 2 cm in diameter. Case 2, a 3-year-old girl complained with a painless tumor of the knee, measuring approximately 1.5 cm in diameter. Microscopically, the tumor had a clear boundary and showed multinodular growth. The tumor was mainly composed of spindle cells arranged in long intersecting fascicles associated with thin, slit-like or branching ectatic vessels, focally forming hemangiopericytoma-like appearance. The tumor cells were abundant, but there was no obvious atypia. Mitotic figures (3-4/10 HPF) were noted. H-caldesmon and SMA were positive in both cases. Case 1 showed diffuse and strong positivity for Desmin, and focally for CKpan. Ki-67 proliferation index was 20% and 30%, respectively. FISH displayed NCOA2 gene translocation in case 1 and the RELA gene translocation in case 2. NGS detected the SRF-NCOA2 gene fusion in case 1 and the SRF-RELA gene fusion in case 2. Both patients underwent local excisions. During the follow-up of 5-14 months, case 1 had no local recurrence, while case 2 developed local recurrence 1 year post operatively. Conclusions: SRF-rearranged cellular perivascular myoid tumor is a novel variant of perivascular cell tumor, which tends to occur in children and adolescents. The tumor forms a broad morphologic spectrum ranging from a pericytic pattern to a myoid pattern, and include hybrid tumors with a mixture of pericytic and myoid patterns. Due to its diffuse hypercellularity and increased mitotic figures and smooth muscle-like immunophenotype, the tumor is easy to be misdiagnosed as myogenic sarcomas. The tumor usually pursues a benign clinical course and rare cases may locally recur.

Droplet digital PCR-based EGFR mutation detection with an internal quality control index to determine the quality of DNA.

In clinical translational research and molecular in vitro diagnostics, a major challenge in the detection of genetic mutations is overcoming artefactual results caused by the low-quality of formalin-fixed paraffin-embedded tissue (FFPET)-derived DNA (FFPET-DNA). Here, we propose the use of an 'internal quality control (iQC) index' as a criterion for judging the minimum quality of DNA for PCR-based analyses. In a pre-clinical study comparing the results from droplet digital PCR-based EGFR mutation test (ddEGFR test) and qPCR-based EGFR mutation test (cobas EGFR test), iQC index ≥ 0.5 (iQC copies ≥ 500, using 3.3 ng of FFPET-DNA [1,000 genome equivalents]) was established, indicating that more than half of the input DNA was amplifiable. Using this criterion, we conducted a retrospective comparative clinical study of the ddEGFR and cobas EGFR tests for the detection of EGFR mutations in non-small cell lung cancer (NSCLC) FFPET-DNA samples. Compared with the cobas EGFR test, the ddEGFR test exhibited superior analytical performance and equivalent or higher clinical performance. Furthermore, iQC index is a reliable indicator of the quality of FFPET-DNA and could be used to prevent incorrect diagnoses arising from low-quality samples.

Regulation of diacylglycerol acyltransferase 2 protein stability by gp78-associated endoplasmic-reticulum-associated degradation.

Triacylglycerol (TG) is the major form of stored energy in eukaryotic organisms and is synthesized by diacylglycerol acyltransferase (DGAT) in the endoplasmic reticulum (ER). DGAT2, one of the two DGAT enzymes, is barely detectable in cells, even though its mRNA transcripts are maintained at considerable levels. However, little is known about how DGAT2 expression is altered by protein stability. DGAT2 was highly unstable in cells and was rapidly degraded by proteasomes in an ubiquitin-dependent manner. Deletion mutation analysis identified transmembrane domain 1 (TMD1) as a protein degradation signal. TMD1 is also important for ER localization of DGAT2. Moreover, DGAT2 interacted with p97/VCP, a crucial component of the ER-associated degradation (ERAD) pathway, and polyubiquitinated DGAT2 accumulated following treatment with an ERAD inhibitor. Furthermore, gp78, an E3 ligase involved in ERAD, regulates the degradation of DGAT2 through direct interactions and ubiquitination. Consequently, the stabilization of DGAT2 increased the number of lipid droplets in hepatic cells. Therefore, DGAT2 is regulated by gp78-associated ERAD at the post-translational level.

A study on the factors affecting the use of the POGS-Nationwide Statistics System (PNSS) among Philippine Obstetrical and Gynecological Society (POGS)-accredited institutions in 2016 / Philippine Journal of Obstetrics and Gynecology

Objective@#To evaluate the factors affecting the use of the POGS PNSS across institutions. @*Basic Procedure@#A cross-sectional study was done by the POGS Committee on Nationwide Statistics from November 2017 to April 2018. Eight representative institutions were chosen from POGS accredited institutions for service and training from the NCR, Luzon, Visayas, and Mindanao, based on the classifications of I. Complete Data, II. Incomplete Data, III. Wrong Format, and IV. No Data. Hospitals with the most number of admissions under each category were chosen. Interviews with chairpersons of the departments, Focus Group Discussions (FGD) with the OB-GYN residents, and actual direct observations of how data were encoded in the PNSS were done.@*Results@#All the chairpersons and Ob-Gyn residents of the selected institutions were aware of the importance of the PNSS especially in generating vital nationwide statistics like Maternal Mortality and Morbidity Rates. They had several pertinent suggestions on how improvement of the PNSS, like harmonization of classification of diseases with PHIC and ICD codes, and to include other co-morbidities in the system.@*Conclusion@#The factors affecting the use of POGS-PNSS in 2016 include: good attitude and compliance among POGSaccredited institutions, but there is a need to address multiple diagnosis including medical co-morbidities. Final diagnosis also needs to be PHIC-compliant, and there is a need to address the Data Privacy Act with the use of eMR (electronic medical records).

Evaluation on the effectiveness for respondent-driven sampling method among men who have sex with men in Beijing, 2017 / 中华流行病学杂志

Objective: To evaluate the randomness and representativeness of respondent- driven sampling (RDS) tool in conducting the investigation in MSM population, in Beijing, 2017. Methods: RDS tool was used to recruit MSM population for a face-to-face interview with structured questionnaire and serological tests. Results: A total of 600 MSM people were sampled and interviewed. The median number of personal network of seeds was 10, which was higher than other MSM people recruited. The numbers of recruitments by wave presented a skewed positive distribution and the highest number was in the fourth wave. It was also dramatically varied from different seeds. Three seeds had the longest chains and had recruited 184, 113 and 92 MSM people, respectively. In contrast, five seeds recruited less than 10 MSM people. Two college students were the most non-generative seeds and each recruited only 1 MSM person. After five to nine waves of sampling, the major demographic characteristics reached equilibrium. Both convergence and bottleneck plots of major demographic characteristics reached convergence, although the plots on marriage and education did not. The homophiles of characteristics were all closed to 1, except for education. The HIV positive rate appeared as 7.9% (95%CI: 4.4%-11.4%) . Conclusions: Results from this study showed that RDS could be used as a feasible sampling method for the study on MSM population with major demographic characteristics reached equilibrium. The process of recruitment appeared controllable and reasonable, showing that this could represent the MSM population in Beijing, in some degree.

Epidemiological characteristics of Coxsackie virus A16 caused hand foot and mouth disease cases in Guangdong province, 2012-2016 / 中华流行病学杂志

Objective: To analyze the epidemiological characteristics of hand foot and mouth disease (HFMD) cases caused by Coxsackie virus A16 (Cox A16) in Guangdong province from 2012 to 2016. Methods: The data of mild HFMD cases caused by Cox A16 were collected from 8 sentinel hospitals in 8 prefecture-level cities in Guangdong to estimate Cox A16 infection status and its population and time distribution characteristics. Results: (1) The highest estimated incidence of Cox A16 infection was in 2014 (113.0/100 000), followed by 2016 (86.4/100 000) and 2012 (79.1/100 000), while the estimated incidence was lower in 2015 (29.0/100 000) and 2013 (28.8/100 000). (2) Cox A16 was confirmed to be the predominant pathogen causing HFMD outbreaks (54.6%, 89/163). The number of outbreaks in the year with high incidence (28 outbreaks) was 11.2 times higher than that in the year with low incidence (2.5 outbreaks). (3) Across all age groups, the annual estimated incidence of Cox A16 infection decreased with age (trend χ(2)=853 905.63, P<0.01). The incidence was highest in age group 1 year (1 449.2/100 000), followed by that in age group 3 years (1 097.0/100 000), in age group 2 years (1 083.5/100 000), in age group 4 years (687.8/100 000) and in age group 0 year (604.9/100 000). Among the age groups <12 months, the estimated incidence increased with age (trend χ(2)=5 541.77, P<0.01), which was highest in age group 11-months (2 105.1/100 000), followed by that in age groups 10-months (1 448.6/100 000), 9-months (938.3/100 000), 8-months (703.3/100 000) and 6-months (664.6/100 000). (4) The annual incidence peak was during May (143.9/100 000)-June (131.5/100 000). Conclusion: The prevalence of Cox A16 infection differed with year in Guangdong during 2012-2016. When the incidence of Cox A16 infection was high, more outbreaks occurred. The prevalence occurred mainly in nurseries and kindergartens from May to June each year. Children aged 0-4 years were the high risk group for Cox A16 infection, children aged 6-11 months were at high risk for Cox A16 infection.

Epidemiological characteristics of outbreaks of norovirus-GⅡ.2, GⅡ.17 and GⅡ.4/Sydney in Guangdong province, 2013-2017 / 中华流行病学杂志

Objective: To understand the epidemiologic characteristics of outbreaks, caused by norovirus-GⅡ.2、GⅡ.17 and GⅡ.4/Sydney in Guangdong Province from 2013 to 2017 and to provide scientific evidence for epidemic prevention and control. Methods: Incidence data of norovirus outbreaks in Guangdong from January 1(st) 2013 to November 30(th) 2017 were collected from Public Health Emergency Management Information System. RT-PCR was performed for every case of each outbreak to detect norovirus nucleic acid and gene sequencing was conducted to identify the genotype of norovirus. Characteristics of norovirus GⅡ.2, GⅡ.17 and GⅡ.4/Sydney outbreaks were analyzed. Directly standardized method was used to calculate the proportion of symtoms as diarrhea and vomitting. Results: From January 1(st) 2013 to November 30(th) 2017, a total of 167 norovirus outbreaks were reported in Guangdong, and 115 outbreaks were caused by norovirus GⅡ.2, GⅡ.17 and GⅡ.4/Sydney respectively. The outbreaks caused by norovirus GⅡ.2 accounted for 39.68% (25/63) in primary schools, 28.57% (18/63) in child care settings, 25.40% (16/63) in middle schools and 6.35% (4/63) in universities. Outbreaks caused by norovirus GⅡ.17 accounted for 41.03% (16/39) in middle schools, 20.51% (8/39) at workplaces, 15.38% (6/39) in primary schools, 12.82% (5/39) in universities, 5.13% (2/39) in communities and child care settings respectively. The outbreaks caused by norovirus GⅡ.4/Sydney accounted for 53.85% (7/13) in universities, 15.38% (2/13) in child care settings and at workplaces respectively, 7.69%(1/13) in primary schools and middle schools respectively. The outbreaks caused by norovirus GⅡ.2 had 77.78% (49/63) of contact transmission, 17.46% (11/63) of food-borne transmission. The outbreaks caused by norovirus GⅡ.17 showed 53.85% (21/39) of food-borne transmission, 15.38% (6/39) of contract transmission, 12.82% (5/39) of water-borne transmission. The outbreaks caused by norovirus GⅡ.4/Sydney had 53.85% (7/13) of food-borne transmission, 38.46% (5/13) of the contact transmission. In terms of the clinical manifestations, the standardized proportion of vomit was 73.76% and the proportion of diarrhea was 42.85% in cases infected with norovirus GⅡ.2, the proportion of standardized of vomit was 76.37% and the proportion of diarrhea was 51.40% in cases infected with norovirus GⅡ.17, with the standardized proportion of vomit was 54.10% and the proportion of diarrhea was 55.95% in cases infected with norovirus GⅡ.4/Sydney. Conclusions: The outbreaks caused by norovirus GⅡ.2 through contact transmission mainly occurred in primary schools, child care settings and middle schools. The outbreaks caused by norovirus GⅡ.17 through food-borne transmission mainly occurred in middle schools and at workplaces. The outbreaks caused by norovirus GⅡ.4/Sydney food-borne transmission and contact mainly occurred in universities.

Recent transmission of pulmonary tuberculosis and its influencing factors in Jing'an district, Shanghai, 2010-2015 / 中华流行病学杂志

Objective: To understand the recent transmission of Mycobacterium tuberculosis (MTB), and to identify the influencing factors of recent transmission among pulmonary tuberculosis (TB) patients in Jing'an district, Shanghai. Methods: The genotypes and drug resistances of MTB isolated from TB patients registered in the TB designated hospitals in Jing'an district during 2010-2015 were analyzed through 12-loci Mycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-VNTR)(QUB11b, QUB18, Mtub21, Miru26, QUB26, Mtub04, Miru31, Miru40, VNTR2372, VNTR3820, 3232, 4120), and tested for drug susceptibility as well. With the results of field epidemiological investigation, univariate and multivariate analyses were performed to analyze the distribution of the clusters and influencing factors on recent transmission. Results: This study enrolled 80 TB patients, 23 (28.75%) had a resistance to at least one anti-TB drug, and the prevalence of multidrug-resistant tuberculosis (MDR-TB) was 16.25%. A total of 65 genotypes were identified with 58 (72.50%, 58/80) being unique and 7 clusters with 2-10 isolated in each cluster. The proportion of clustering was 27.50% (22/80). Results from the multivariate analysis revealed that multidrug- resistance (OR=35.799, 95%CI: 4.239-302.346) and having comorbidity with TB (OR=7.695, 95%CI: 1.421-41.658) were independently associated with the clustering, which suggesting a recent transmission. The field investigation to the clustered cases proved that the patients in two clusters had epidemiological links, one was between family members, and the other contained 10 MDR-TB patients with 9 knowing each other which have a definite connection and 1 having the possible connection with them. Conclusion: Recent transmission of tuberculosis happened among TB patients in Jing'an district, with high risks among the MDR-TB patients.

Theoretical calculation and experimental study of membrane thickness of alginate-(poly-L-lysine)-alginate microcapsules / 生物医学工程学杂志

Alginate-(Poly-L-Lysine)-Alginate(APA) microcapsules were prepared by Electrostatic Droplet Generator(EDG) technique and the thickness of microcapsule membrane, which was composed by polyelectrolyte complex, were studied in this paper. The theoretical formula was given for the measurement of membrane thickness of APA microcapsules by element analysis of membrane and calculation. The membrane thickness was 7-10 microns by theoretical calculation. On the other hand, the thickness of membrane was measured by SEM and optical microscopy and the results were 7 microns and 12 microns, respectively. The results showed that theoretical calculation is in good accordance with experimental determoination of mermbrane thickness and the membrane thickness of APA microcapsule is about 7-10 microns. The optical microscopy is an easy way to measure membrane thickness.