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PurposeFractionated radiotherapy as well as concomitant and adjuvant chemotherapy such as temozolomide for postoperative high-grade glioma (HGG) patients improves progression-free survival and overall survival. Multiple factors such as chemotherapy, radiotherapy, tumor grade, residual tumor volume, and genetic modifications might play a role in the formation of cognitive impairment. The risk factors of cognitive impairment in postoperative patients with HGG receiving radiotherapy and chemotherapy remains a concern in this population. The purpose of this study was to identify risk factors for cognitive impairment in patients of postoperative HGG.Materials and MethodsA total of 229 patients with HGG who underwent surgery were analyzed. Cognitive impairment was defined as a decrease of Cognitive Assessment Montreal (MoCA)’s score in at least two cognitive domains or any MoCA’s score of less than 26 points at the time of study compared with baseline level. Multiple potential risk factors including methylated status of the O6-methylguanine-DNA methyltransferase (MGMT) promoter, glioma World Health Organization (WHO) grade, residual tumor volume, education, and sex were analyzed. Cox univariate and multivariate regression analysis was used to detect the significant risk factors for cognitive impairment.ResultsAt the end of follow-up among the 229 patients, 147 patients (67%) developed cognitive impairment. 82 patients (36%) remained in normal cognitive condition. In multivariate analysis, unmethylated MGMT promoter (hazard ratio [HR], 1.679; 95% confidence interval [CI], 1.212 to 2.326; p=0.002), glioblastoma (HR, 1.550; 95% CI, 1.117 to 2.149; p=0.009), and residual tumor volume > 5.58 cm3 (HR, 1.454; 95% CI, 1.047 to 2.020; p=0.026) were independent risk factors for cognitive impairment.ConclusionMethylated status of the MGMT promoter, glioma WHO grade, and residual tumor volume might be risk factors for the cognitive impairment in postoperative patients with HGG.

Prostaglandin E2 receptor EP1 (PGE2/EP1) promotes diabetic renal injury, and EP1 receptor deletion improves hyperfiltration, albuminuria, and fibrosis. The role of EP1 receptors in hypertensive kidney disease (HKD) remains controversial. We examined the contribution of EP1 receptors to HKD. EP1 null (EP1-/-) mice were bred with hypertensive TTRhRen mice (Htn) to evaluate kidney function and injury at 24 weeks. EP1 deletion had no effect on elevation of systolic blood pressure in Htn mice (HtnEP1-/-) but resulted in pronounced albuminuria and reduced FITC-inulin clearance, compared with Htn or wild-type (WT) mice. Ultrastructural injury to podocytes and glomerular endothelium was prominent in HtnEP1-/- mice; including widened subendothelial space, subendothelial lucent zones and focal lifting of endothelium from basement membrane, with focal subendothelial cell debris. Cortex COX2 mRNA was increased by EP1 deletion. Glomerular EP3 mRNA was reduced by EP1 deletion, and EP4 by Htn and EP1 deletion. In WT mice, PGE2 increased chloride reabsorption via EP1 in isolated perfused thick ascending limb (TAL), but PGE2 or EP1 deletion did not affect vasopressin-mediated chloride reabsorption. In WT and Htn mouse inner medullary collecting duct (IMCD), PGE2 inhibited vasopressin-water transport, but not in EP1-/- or HtnEP1-/- mice. Overall, EP1 mediated TAL and IMCD transport in response to PGE2 is unaltered in Htn, and EP1 is protective in HKD.

Background: We determined to assess the HDAC8 expression in esophageal squamous cell carcinoma (ESCC) patients and prognostic potential though there is only a little research contribution regarding HDAC8 to tumorigenesis of ESCC. Methods: The immunohistochemical expression of HDAC8 was investigated on tissue microarrays (TMAs) from 110 patients with esophageal squamous cell carcinoma.The nuclear staining intensity is calculated by the immune reactivity score ranging from (0-12) and divides them into two groups: no expression group and overexpression. Results: The median follow-up duration was 70 months. Highly regulated HDAC8 protein significantly predicted decreased 5-year overall survival (p = 0.001) and progression-free survival (p = 0.001) demonstrated by the log-rank test. Furthermore, HDAC8 protein acts as an independent prognostic factor for overall survival and progression-free survival, which determined after multivariate analyses. By Receiver operating characteristics (ROC) analysis, the value of HDAC8 was (0.63±0.54,p=0.04) according to advance cancer stage and (0.59±0.06,p=0.04) according to the lymph node status found in the Area under the curve (AUC). Conclusion: HDAC8 protein is highly regulated in ESCC tissues and potential prognostic indicator for diagnosing patients with decreased survival period.

Rhoifolin (ROF) is a main effective component in Citrus grandis 'Tomentosa'. ROF has a potential anti-inflammatory activity, but its specific effects and mechanisms have not been studied. This study investigated the anti-inflammatory activity of ROF and searched for its possible molecular mechanisms. A mouse model of acute inflammation was induced by lipopolysaccharide, and the effects of ROF on pathological damages of the lung and liver were observed. Carrageenan-induced paw edema rat model was used to evaluate the effect of ROF on the volume of swelling paw. In LPS-induced RAW264.7 macrophages, the expression levels of pro-inflammatory cytokines IL-1β, IL-6, and TNF-α were measured using ELISA. Real-time PCR was used to measure the mRNA levels of iNOS and CCL2. Western blot was used to detect the activation of IκBα and IKKβ in NF-κB signaling pathways. The results showed that ROF accelerated the recoveries of liver and lung tissue damages in acute inflammation mice and inhibited carrageenan-induced paw edema in rats; in addition, ROF significantly suppressed the secretion of TNF-α, IL-1β, and IL-6 in the serum of rats and mouse model. In LPS-induced RAW264.7 cells, 100 μmol/L ROF enhanced cell viability and suppressed the production of TNF-α, IL-6, and IL-1β significantly. ROF also decreased the mRNA expression of iNOS and CCL2 and inhibited IκBα and IKKβ phosphorylation. In summary, ROF had a potential therapeutic value for inflammation. Our research provided experimental basis for the further development of ROF as an anti-inflammatory drug and for clarifying the anti-inflammatory substance basis of Citrus grandis 'Tomentosa'. Graphical Abstract.

Germline mutations in BRCA1 and BRCA2 are the most penetrating genetic predispositions for breast and ovarian cancer, and their presence is largely ethnic-specific. Comprehensive information about the prevalence and spectrum of BRCA mutations has been collected in European and North American populations. However, similar information is lacking in other populations, including the mainland Chinese population despite its large size of 1.4 billion accounting for one fifth of the world's population. Herein, we performed an extensive literature analysis to collect BRCA variants identified from mainland Chinese familial breast and ovarian cancer patients. We observed 137 distinct BRCA1 variants in 409 of 3,844 and 80 distinct BRCA2 variants in 157 of 3,024 mainland Chinese patients, with an estimated prevalence of 10.6% for BRCA1 and 5.2% for BRCA2. Of these variants, only 40.3% in BRCA1 and 42.5% in BRCA2 are listed in current Breast Cancer Information Core database. We observed higher frequent variation in BRCA1 exons 11A, 11C, 11D, and 24 and BRCA2 exon 10 in Chinese patients than in the patients of other populations. The most common pathogenic variant in BRCA1 wasc.981_982delAT in exon 11A, and in BRCA2 c.3195_3198delTAAT in exon 11B and c.5576_5579delTTAA in exon 11E; the most common novel variant in BRCA1 was c.919A>G in exon 10A, and in BRCA2 c.7142delC in exon 14. None of the variants overlap with the founder mutations in other populations. Our analysis indicates that the prevalence of BRCA variation in mainland Chinese familial breast and ovarian cancer patients is at a level similar to but the spectrum is substantially different from the ones of other populations.

When recombinant human (rh) thyroid-stimulating hormone (TSH) is administered to thyroid cancer survivors, an acute extra-thyroidal effect raises pro-inflammatory cytokines and activates platelets. Thymic stromal lymphopoietin (TSLP) is a cytokine recently implicated in platelet activation. Our aim was to measure platelet microparticle levels after rhTSH stimulation in vivo, and to investigate TSLP expression in TSH-stimulated human adipocytes in culture. Blood samples for total and platelet microparticle analysis were obtained from thyroid cancer survivors before (day 1) and after rhTSH administration (day 5). Adipocytes, differentiated from stromal preadipocytes isolated from adipose tissue from surgical patients, were stimulated with TSH. TSLP mRNA expression, protein expression, and protein release into the adipocyte medium were measured. The level of platelet microparticles in thyroid cancer patients rose 5-fold after rhTSH stimulation. TSH upregulated TSLP mRNA expression in adipocytes in culture through a pathway that was inhibited by 66% by H89, a protein kinase A inhibitor. TSLP protein expression rose in response to TSH, and TSH-stimulated TSLP release into the medium was completely blocked by dexamethasone. In conclusion, TSLP is a novel TSH-responsive adipokine. Future studies will be needed to address the potential role of adipocyte-derived TSLP and whether it is linked to TSH-dependent platelet activation.

Angiotensin-converting enzyme 2 (ACE2) is a recently described member of the renin-angiotensin system that hydrolyzes angiotensin (Ang) II to Ang-(1-7), and may thereby protect against cardiovascular and renal diseases. ACE2 is a type 1 integral membrane protein and contains a catalytically active ectodomain that can be shed from the cell surface into the extracellular space, via cleavage by a disintegrin and metalloproteinase-17 (ADAM-17). ACE2 enzymatic activity and protein can be detected in biological fluids, including urine, plasma, and conditioned cell culture media. We present a detailed method for measurement of ACE2 activity in biological fluids, using hydrolysis of an intramolecularly quenched fluorogenic ACE2 substrate, in the absence or presence of the ACE2 inhibitors MLN-4760 or DX600. Recombinant human or mouse ACE2 is used to generate standard curves for this assay, with ACE2 detection ranging from 1.56 to 50 ng/ml. While MLN-4760 potently inhibits the activity of both human and mouse ACE2, DX600 (linear form) only effectively blocks human ACE2 activity in this assay. In biological samples of human and mouse urine, cell culture medium from mouse proximal tubular cells, and mouse plasma, the mean intra- and inter-assay coefficients of variation (CVs) of the assay range from 1.43 to 4.39 %, and from 7.01 to 13.17 %, respectively. We present data on the time and substrate concentration dependence of the assay, and show that exogenous D -glucose, creatinine, urea, and albumin do not interfere with its performance. In biological fluids, this assay is a simple and reliable method to study the role of ACE2 and its shed fragments in cardiovascular and renal diseases.

Assessment of urinary extracellular vesicles including exosomes and microparticles (MPs) is an emerging approach for non-invasive detection of renal injury. We have previously reported that podocyte-derived MPs are increased in diabetic mice in advance of albuminuria. Here, we hypothesised that type 1 diabetes and acute hyperglycaemia would increase urinary podocyte MP levels in uncomplicated diabetes.In this post hoc exploratory analysis, we examined archived urine samples from normoalbuminuric patients with uncomplicated type 1 diabetes studied under clamped euglycaemia and hyperglycaemia and compared with healthy controls. Urinary vesicles were assessed by electron microscopy and nanoparticle tracking while podocyte MPs were assessed by flow cytometry.Neither vesicle size nor total number were significantly altered in type 1 diabetes or acute hyperglycaemia. By contrast, urinary podocyte MP levels were higher in type 1 diabetes (0.47 [0.00-3.42] MPs/μmol creatinine [Cr]) compared with healthy controls (0.00 [0.00-0.00] MPs/μmol Cr, p 0.05) and increased under hyperglycaemic clamp (0.36 [0.00-4.15] MPs/μmol Cr during euglycaemia vs 2.70 [0.00-15.91] MPs/μmol Cr during hyperglycaemia, p 0.05). Levels of urinary albumin to creatinine ratio and nephrin (surrogates of podocyte injury) were unchanged by type 1 diabetes or acute hyperglycaemia.Taken together, our data show that urinary podocyte MP levels are higher in patients with type 1 diabetes in advance of changes in other biomarkers (albuminuria, nephrin). Examination of podocyte MPs may serve as an early biomarker of glomerular injury in uncomplicated type 1 diabetes.

Core promoter controls the initiation of transcription. Core promoter sequence change can disrupt transcriptional regulation, lead to impairment of gene expression and ultimately diseases. Therefore, comprehensive characterization of core promoters is essential to understand normal and abnormal gene expression in biomedical studies. Here we report the development of EVDC (Exome-based Variant Detection in Core promoters) method for genome-scale analysis of core-promoter sequence variation. This method is based on the fact that exome sequences contain the sequences not only from coding exons but also from non-coding region including core promoters generated by random fragmentation in exome sequencing process. Using exome data from three cell types of CD4+ T cells, CD19+ B cells and neutrophils of a single individual, we characterized the features of core promoter-mapped exome sequences, and analysed core-promoter variation in this individual genome. We also compared the core promoters between YRI (Yoruba in Ibadan, Nigeria) and the CEU (Utah residents of European decedent) populations using the exome data generated by the 1000 Genome project, and observed much higher variation in YRI population than in CEU population. Our study demonstrates that the EVDC method provides a simple but powerful means for genome-wile de novo characterization of core promoter sequence variation.

Angiotensin-converting enzyme 2 (ACE2) degrades angiotensin (Ang) II to Ang-(1-7), and protects against diabetic renal injury. Soluble ACE2 fragments are shed from the proximal tubule, and appear at high levels in the urine with diabetes. High glucose-induced shedding of ACE2 from proximal tubular cells is mediated by the enzyme "a disintegrin and metalloproteinase-17″ (ADAM17). Here, we investigated the mechanism for constitutive shedding of ACE2. Mouse proximal tubular cells were cultured and ACE2 shedding into the media was assessed by enzyme activity assay and immunoblot analysis. Cells were incubated with pharmacologic inhibitors, or transfected with silencing (si) RNA. Incubation of proximal tubular cells with increasing concentrations of D-glucose stimulated ACE2 shedding, which peaked at 16 mM, while L-glucose (osmotic control) had no effect on shedding. In cells maintained in 7.8 mM D-glucose, ACE2 shedding was significantly inhibited by the pan-protein kinase C (PKC) competitive inhibitor sotrastaurin, but not by an inhibitor of ADAM17. Incubation of cells with the PKC-α and -β1-specific inhibitor Go6976, the PKC β1 and β2-specific inhibitor ruboxistaurin, inhibitors of matrix metalloproteinases-2,-8, and -9, or an inhibitor of ADAM10 (GI250423X) had no effect on basal ACE2 shedding. By contrast, the PKC-δ inhibitor rottlerin significantly inhibited both constitutive and high glucose-induced ACE2 shedding. Transfection of cells with siRNA directed against PKC-δ reduced ACE2 shedding by 20%, while knockdown of PKC-ε was without effect. These results indicate that constitutive shedding of ACE2 from proximal tubular cells is mediated by PKC-δ, which is also linked to high glucose-induced shedding. Targeting PKC-δ may preserve membrane-bound ACE2 in proximal tubule in disease states and diminish Ang II-stimulated adverse signaling.

Angiotensin-converting enzyme 2 (ACE2) degrades angiotensin II to angiotensin-(1–7) and is expressed in podocytes. Here we overexpressed ACE2 in podocytes in experimental diabetic nephropathy using transgenic methods where a nephrin promoter drove the expression of human ACE2. Glomeruli from these mice had significantly increased mRNA, protein, and activity of ACE2 compared to wild-type mice. Male mice were treated with streptozotocin to induce diabetes. After 16 weeks, there was no significant difference in plasma glucose levels between wild-type and transgenic diabetic mice. Urinary albumin was significantly increased in wild-type diabetic mice at 4 weeks, whereas albuminuria in transgenic diabetic mice did not differ from wild-type nondiabetic mice. However, this effect was transient and by 16 weeks both transgenic and nontransgenic diabetic mice had similar rates of proteinuria. Compared to wild-type diabetic mice, transgenic diabetic mice had an attenuated increase in mesangial area, decreased glomerular area, and a blunted decrease in nephrin expression. Podocyte numbers decreased in wild-type diabetic mice at 16 weeks, but were unaffected in transgenic diabetic mice. At 8 weeks, kidney cortical expression of transforming growth factor-β1 was significantly inhibited in transgenic diabetic mice as compared to wild-type diabetic mice. Thus, the podocyte-specific overexpression of human ACE2 transiently attenuates the development of diabetic nephropathy.

Background Genetic predisposition is the primary risk factor for familial breast cancer. For the majority of familial breast cancer, however, the genetic predispositions remain unknown. All newly identified predispositions occur rarely in disease population, and the unknown genetic predispositions are estimated to reach up to total thousands. Family unit is the basic structure of genetics. Because it is an autosomal dominant disease, individuals with a history of familial breast cancer must carry the same genetic predisposition across generations. Therefore, focusing on the cases in lineages of familial breast cancer, rather than pooled cases in disease population, is expected to provide high probability to identify the genetic predisposition for each family. Methods In this study, we tested genetic predispositions by analyzing the family-specific variants in familial breast cancer. Using exome sequencing, we analyzed three families and 22 probands with BRCAx (BRCA-negative) familial breast cancer. Results We observed the presence of family-specific, novel, deleterious germline variants in each family. Of the germline variants identified, many were shared between the disease-affected family members of the same family but not found in different families, which have their own specific variants. Certain variants are putative deleterious genetic predispositions damaging functionally important genes involved in DNA replication and damaging repair, tumor suppression, signal transduction, and phosphorylation. Conclusions Our study demonstrates that the predispositions for many BRCAx familial breast cancer families can lie in each disease family. The application of a family-focused approach has the potential to detect many new predispositions.

Background BRCA1 plays an essential role in maintaining genome stability. Inherited BRCA1 germline mutation (BRCA1+) is a determined genetic predisposition leading to high risk of breast cancer. While BRCA1+ induces breast cancer by causing genome instability, most of the knowledge is known about somatic genome instability in breast cancer cells but not germline genome instability. Methods Using the exome-sequencing method, we analyzed the genomes of blood cells in a typical BRCA1+ breast cancer family with an exon 13-duplicated founder mutation, including six breast cancer-affected and two breast cancer unaffected members. Results We identified 23 deleterious mutations in the breast cancer-affected family members, which are absent in the unaffected members. Multiple mutations damaged functionally important and breast cancer-related genes, including transcriptional factor BPTF and FOXP1, ubiquitin ligase CUL4B, phosphorylase kinase PHKG2, and nuclear receptor activator SRA1. Analysis of the mutations between the mothers and daughters shows that most mutations were germline mutation inherited from the ancestor(s) while only a few were somatic mutation generated de novo. Conclusion Our study indicates that BRCA1+ can cause genome instability with both germline and somatic mutations in non-breast cells.

Angiotensin-converting enzyme 2 (ACE2) is expressed in the kidney and may be a renoprotective enzyme, since it converts angiotensin (Ang) II to Ang-(1-7). ACE2 has been detected in urine from patients with chronic kidney disease. We measured urinary ACE2 activity and protein levels in renal transplant patients (age 54 yrs, 65% male, 38% diabetes, n = 100) and healthy controls (age 45 yrs, 26% male, n = 50), and determined factors associated with elevated urinary ACE2 in the patients. Urine from transplant subjects was also assayed for ACE mRNA and protein. No subjects were taking inhibitors of the renin-angiotensin system. Urinary ACE2 levels were significantly higher in transplant patients compared to controls (p = 0.003 for ACE2 activity, and p≤0.001 for ACE2 protein by ELISA or western analysis). Transplant patients with diabetes mellitus had significantly increased urinary ACE2 activity and protein levels compared to non-diabetics (p

Genetic aberrations contribute to acute myeloid leukemia (AML). However, half of AML cases do not contain the well-known aberrations detectable mostly by cytogenetic analysis, and these cases are classified as normal karyotype AML. Different outcomes of normal karyotype AML suggest that this subgroup of AML could be genetically heterogeneous. But lack of genetic markers makes it difficult to further study this subgroup of AML. Using paired-end RNAseq method, we performed a transcriptome analysis in 45 AML cases including 29 normal karyotype AML, 8 abnormal karyotype AML and 8 AML without karyotype informaiton. Our study identified 134 fusion transcripts, all of which were formed between the partner genes adjacent in the same chromosome and distributed at different frequencies in the AML cases. Seven fusions are exclusively present in normal karyotype AML, and the rest fusions are shared between the normal karyotype AML and abnormal karyotype AML. CIITA, a master regulator of MHC class II gene expression and truncated in B-cell lymphoma and Hodgkin disease, is found to fuse with DEXI in 48% of normal karyotype AML cases. The fusion transcripts formed between adjacent genes highlight the possibility that certain such fusions could be involved in oncological process in AML, and provide a new source to identify genetic markers for normal karyotype AML.