The association of telomere maintenance and TERT expression with susceptibility to human papillomavirus infection in cervical epithelium.

The role of telomerase reverse transcriptase (TERT) induction and telomere maintenance in carcinogenesis including cervical cancer (CC) pathogenesis has been well established. However, it remains unclear whether they affect infection of high-risk human papillomavirus (hrHPV), an initiating event for CC development. Similarly, genetic variants at the TERT locus are shown to be associated with susceptibility to CC, but it is unclear whether these SNPs modify the risk for cervical HPV infection. Here we show that in CC-derived HeLa cells, TERT overexpression inhibits, while its depletion upregulates expression of Syndecan-1 (SDC-1), a key component for HPV entry receptors. The TCGA cohort of CC analyses reveals an inverse correlation between TERT and SDC-1 expression (R = -0.23, P = 0.001). We further recruited 1330 females (520 non-HPV and 810 hrHPV-infected) without CC or high-grade cervical intraepithelial neoplasia to analyze telomeres in cervical epithelial cells and SNPs at rs2736098, rs2736100 and rs2736108, previously identified TERT SNPs for CC risk. Non-infected females exhibited age-related telomere shortening in cervical epithelial cells and their telomeres were significantly longer than those in hrHPV-infected group (1.31 ± 0.62 vs 1.19 ± 0.48, P < 0.001). There were no differences in rs2736098 and rs2736100 genotypes, but non-infected individuals had significantly a higher C-allele frequency (associated with higher TERT expression) while lower T-allele levels at rs2736108 compared with those in the hrHPV group (P = 0.020). Collectively, appropriate telomere maintenance and TERT expression in normal cervical cells may prevent CC by modulating hrHPV infection predisposition, although they are required for CC development and progression.

Janus hydrogel loaded with a CO2-generating chemical reaction system: Construction, characterization, and application in fruit and vegetable preservation.

In this study, we inserted a dynamic chemical reaction system that can generate CO2 into Janus hydrogel (JH) to develop a multidimensional preservation platform that integrates hygroscopicity, antibacterial activity, and modified atmospheric capacity. The double gel system developed using sodium alginate/trehalose at a 1:1 ratio effectively encapsulated 90% of citric acid. Furthermore, CO2 loss was avoided by separately embedding NaHCO3/cinnamon essential oil and citric acid microcapsules into a gelatin pad to develop JH. Freeze-dried JH exhibited a porous and asymmetric structure, very strongly absorbing moisture, conducting water, and rapidly releasing CO2 and essential oils. Furthermore, when preserving various fruits and vegetables in practical settings, JH provided several preservation effects, including color protection, microbial inhibition, and antioxidant properties. Our study findings broaden the application of JH technology for developing chemical reaction systems, with the resulting JH holding substantial promise for cold chain logistics.

Genomic and transcriptomic analysis of breast cancer identifies novel signatures associated with response to neoadjuvant chemotherapy.

BACKGROUND: Neoadjuvant chemotherapy (NAC) has become a standard treatment strategy for breast cancer (BC). However, owing to the high heterogeneity of these tumors, it is unclear which patient population most likely benefit from NAC. Multi-omics offer an improved approach to uncovering genomic and transcriptomic changes before and after NAC in BC and to identifying molecular features associated with NAC sensitivity. METHODS: We performed whole-exome and RNA sequencing on 233 samples (including matched pre- and post-treatment tumors) from 50 BC patients with rigorously defined responses to NAC and analyzed changes in the multi-omics landscape. Molecular features associated with NAC response were identified and validated in a larger internal, and two external validation cohorts, as well as in vitro experiments. RESULTS: The most frequently altered genes were TP53, TTN, and MUC16 in both pre- and post-treatment tumors. In comparison with pre-treatment tumors, there was a significant decrease in C > A transversion mutations in post-treatment tumors (P = 0.020). NAC significantly decreased the mutation rate (P = 0.006) of the DNA repair pathway and gene expression levels (FDR = 0.007) in this pathway. NAC also significantly changed the expression level of immune checkpoint genes and the abundance of tumor-infiltrating immune and stroma cells, including B cells, activated dendritic cells, γδT cells, M2 macrophages and endothelial cells. Furthermore, there was a higher rate of C > T substitutions in NAC nonresponsive tumors than responsive ones, especially when the substitution site was flanked by C and G. Importantly, there was a unique amplified region at 8p11.23 (containing ADGRA2 and ADRB3) and a deleted region at 3p13 (harboring FOXP1) in NAC nonresponsive and responsive tumors, respectively. Particularly, the CDKAL1 missense variant P409L (p.Pro409Leu, c.1226C > T) decreased BC cell sensitivity to docetaxel, and ADGRA2 or ADRB3 gene amplifications were associated with worse NAC response and poor prognosis in BC patients. CONCLUSIONS: Our study has revealed genomic and transcriptomic landscape changes following NAC in BC, and identified novel biomarkers (CDKAL1P409L, ADGRA2 and ADRB3) underlying chemotherapy resistance and poor prognosis, which could guide the development of personalized treatments for BC.

Vitamin D Promotes Ferroptosis in Colorectal Cancer Stem Cells via SLC7A11 Downregulation.

Colorectal cancer stem cells (CCSCs) play important roles in the prognosis, chemoresistance, and treatment failure of colorectal cancer (CRC). Ferroptosis is an effective treatment for CCSCs. Vitamin D (VD) reportedly inhibits colon cancer cell proliferation. However, information on the relationship between VD and ferroptosis in CCSCs is not well documented. In this study, we aimed to understand the effect of VD on ferroptosis in CCSCs. To this end, we treated CCSCs with different concentrations of VD and performed spheroid formation assay and transmission electron microscopy and determined cysteine (Cys), glutathione (GSH), and reactive oxygen species (ROS) levels. Furthermore, functional experiments, western blotting, and qRT-PCR were performed to explore the downstream molecular mechanisms of VD in vitro and in vivo. Results showed that VD treatment significantly inhibited the proliferation of CCSCs and reduced the number of tumour spheroids in vitro. Further evaluations showed that the VD-treated CCSCs exhibited significantly higher ROS levels and lower levels of Cys and GSH as well as thickened mitochondrial membranes. Furthermore, the mitochondria in CCSCs were narrowed and ruptured after VD treatment. These results indicated that VD treatment significantly induced ferroptosis in CCSCs. Further exploration showed that SLC7A11 overexpression significantly attenuated VD-induced ferroptosis in vitro and in vivo. Hence, we concluded that VD induces ferroptosis in CCSCs by downregulating SLC7A11 in vitro and in vivo. These results provide new evidence for the therapeutic use of VD in treating CRC and new insights into VD-induced ferroptosis in CCSCs.

[Expression and function of vascular endothelial growth factor and angiopoietins in rat brain after cerebral ischemia].

OBJECTIVE: To examine the temporal and spatial expression of vascular endothelial growth factor (VEGF) and angiopoietins (Ang) in rat brain after cerebral ischemia, and to elucidate the roles they played in angiogenesis and vascular permeability. METHODS: Rats were subjected to either middle cerebral artery occlusion (MCAO) or sham operation. Reverse transcriptase-polymerase chain reaction, Western blotting, and immunohistochemistry were used to detect the expression of VEGF, Ang-1 and Ang-2 at different time points after ischemia. CD31 was used to label endothelial cells after MCAO. Vascular permeability was determined by Evans blue. RESULTS: VEGF was markedly increased at 2 h, had an initial peak at 12 h (0.7249 ± 0.1933, P < 0.01), and a second peak at 7 days (0.5264 ± 0.1519, P < 0.01). Ang-2 mRNA and protein significantly increased after MCAO, both of them peaked at 12 h (0.6747 ± 0.2416, P < 0.01; 1.1197 ± 0.1780, P < 0.01). In contrast, Ang-1 mRNA and protein gradually decreased after MCAO, respectively reaching a minimum at 3 d (0.3220 ± 0.1427, P < 0.01) and 1 d (0.1298 ± 0.0293, P < 0.01). Changes in the expression of these factors correlated with the progress of angiogenesis and vascular permeability. Evans blue test revealed that the vascular permeability gradually increased, and peaked at day 1 after ischemia [(6.219 ± 0.887) µg/g, P < 0.01]. CONCLUSION: Dynamic temporal changes in VEGF, Ang-1 and Ang-2 expression stimulate the cerebral angiogenesis after focal cerebral ischemia.

CCL20 mediates the anti-tumor effect of vitamin D3 in p38MAPK/NF-κB signaling in colitis-associated carcinogenesis.

Vitamin D3 is beneficial in ameliorating or preventing inflammation and carcinogenesis. CCL20 is a potential therapeutic target in carcinogenesis, which mediates the protective effect of vitamin D or vitamin D analogue in autoimmune and cancer diseases. Here we aim to evaluate whether vitamin D3 plays a protective role in colitis-associated colorectal cancer (CAC) by affecting CCL20 and the molecular mechanism. Administration of azoxymethane (AOM) followed with dextran sulfate sodium (DSS) was used to simulate CAC in mouse. After 5-day DSS treatment, vitamin D3 supplementation was for 9 weeks at 60 IU/g/w. We found that dietary vitamin D3 significantly reduced the tumor number and tumor burden in mouse. In-vivo and -vitro, vitamin D3 reduced the levels of CCL20, phospho-p38 MAPK (p-p38) and phospho-NF-κB p65 (p-p65), and the transcriptional activity of NF-κB. Further studies showed that CCL20 mediated the inhibition of vitamin D3 in p38MAPK-mediated NF-κB signaling in vitro. Taken together, vitamin D3 effectively suppressed colonic carcinogenesis in AOM-DSS mouse model. Downregulation of CCL20 may contribute to the preventive effect of vitamin D3 on NF-κB activity. It may merit further clinical investigation as a therapeutic agent against CAC in humans.

Novel compound variants of the AR and MAP3K1 genes are related to the clinical heterogeneity of androgen insensitivity syndrome.

Androgen insensitivity syndrome (AIS; OMIM 300068) is the most frequent cause of 46, XY disorders of sex development (DSD). However, the correlation between genotype and phenotype has not been determined. We conducted a systematic analysis of the clinical characteristics, hormone levels, ultrasonography data and histopathology of a 46, XY Chinese patient with AIS. The family was followed up for nearly 8 years. We applied whole-exome sequencing (WES) for genetic analysis of the pedigree and performed bioinformatic analysis of the identified variants. Human embryonic kidney 293T/17 (HEK293T/17) cells were transiently transfected with wild-type or mutant AR and MAP3K1 plasmid. Cell lysates were used to analyze androgen receptor (AR) production. A novel hemizygous AR variant (c.2070C>A, p. His690Glu) and a rare heterozygous MAP3K1 variant (c.778C>T, p. Arg260Cys) were identified by WES in the proband and her mother. Bioinformatic analysis predicted these two variants to be pathogenic. Multiple amino acid sequence alignments showed that p. His690 and p. Arg260 are conserved among various species. His690Glu is a mutation that decreased the AR production, whereas the Arg260Cys mutation increased the AR production. The novel compound variants of the AR and MAP3K1 genes also increased the production of AR protein. Thus, the phenotype of the patient may be caused by defects in both the AR and MAP3K1 signaling pathways. Compound variants of the AR and MAP3K1 genes resulted in a specific phenotype in this patient with AIS. WES might reveal genetic variants that explain the heterogeneity of AIS.

Metformin Delays the Development of Atherosclerosis in Type 1 Diabetes Mellitus via the Methylglyoxal Pathway.

INTRODUCTION: The aim of our study was to determine the effect of metformin administration on juvenile type 1 diabetes mellitus and atherosclerosis in apolipoprotein E null (ApoE-/-) mice and to explore the mechanism involved. METHODS: Eighteen male ApoE-/- mice were injected with streptozotocin to induce diabetes (diabetic group) and 18 mice who received no streptozotocin injection were assigned to the control (non-diabetic) group. Six mice in each group were then orally administered metformin, simvastatin, or vehicle, respectively, following which the mice were euthanized and tissue samples collected. RESULTS: Fasting plasma glucose, low-density lipoprotein-cholesterol, and triglyceride concentrations were significantly higher in the three diabetic groups than in the three non-diabetic groups. Plasma N∈-(carboxymethyl)lysine and N∈-(carboxyethyl)lysine concentrations were higher in the diabetic mice than in the non-diabetic mice, but metformin treatment reduced these concentrations more effectively than simvastatin. All three diabetic groups demonstrated obvious arterial plaques, but these were largest in the vehicle-treated diabetic group. The expression of extracellular nitric oxide synthase was highest in the simvastatin-treated non-diabetic group, and in diabetic mice it was higher in the simvastatin-treated group than in the other two groups. No significant expression of AMP-activated protein kinase (AMPK) was measured in the three diabetic groups, but a low level of AMPK expression was detected in the non-diabetic groups. CONCLUSIONS: Metformin can limit the development of atherosclerosis secondary to diabetes in young diabetic mice. A possible mechanism is the removal of methylglyoxal, thereby reducing the formation of advanced glycation endproducts, rather than by lowering the blood glucose level. FUNDING: This work was supported by the National Natural Science Foundation of China (81901106) and Jinan clinical medical science and technology innovation plan (201907002).

The effect of bone marrow mesenchymal stem cell and nano-hydroxyapatite/collagen I/poly-L-lactic acid scaffold implantation on the treatment of avascular necrosis of the femoral head in rabbits.

For avascular necrosis of the femoral head (ANFH), repair and regeneration are difficult because of the edema and high pressure caused by continuous ischemia and hypoxia. Core decompression (CD) is a classic method for treating early ANFH before the collapse of the femoral head; however, its effect is still controversial. To improve the therapeutic effect of CD on ANFH, a novel tissue-engineered bone (TEB) was constructed by combining bone marrow mesenchymal stem cells (BMSCs) with nano-hydroxyapatite/collagen I/poly-L-lactic acid (nHAC/PLA) scaffolds and implanting the TEB into the bone tunnel of CD. Cell attachment was observed by scanning electron microscopy and hematoxylin and eosin staining. The authors' previous studies confirmed that nHAC/PLA is an excellent scaffold material with favorable biocompatibility and no cytotoxicity. A total of 24 New Zealand rabbits with ANFH were randomly divided into three groups, as follows: Group A (n=8), pure CD; group B (n=8), CD+nHAC/PLA; and group C (n=8), CD+BMSCs-nHAC/PLA. The favorable effect of BMSCs-nHAC/PLA on angiogenesis and bone formation in necrotic areas was further evaluated via radiographic and histological analyses. Computerized tomography (CT) scanning and H&E staining showed more capillaries and new osteoid tissue in group C compared with in groups B and A. Micro-CT showed that the new bone coverage rate and implanted material degradation degree were each increased in group C compared with in group B. These results indicate that BMSCs-nHAC/PLA scaffolds may improve the curative effect of CD and provide a strategy for treating ANFH.

Synchronous Neoplastic Lesions In Referred Patients With Colorectal Cancer: A Retrospective Cohort Study.

BACKGROUND: Synchronous neoplastic lesions are usually present in patients with colorectal cancer (CRC) at diagnosis or postoperative follow-up endoscopy. However, few studies have been published about the clinicopathological features of synchronous lesions, especially those of synchronous advanced neoplasia. This study aimed to describe synchronous lesions in patients with CRC because this knowledge may be useful for preventing the development of metachronous cancer. MATERIAL AND METHODS: We retrospectively reviewed 261 primary CRC cases with synchronous lesions referred to our hospital during a 4-year period. Personal history, habits, family history, characteristics of index cancer, and synchronous lesions were assessed. RESULTS: In total, the 261 patients with CRC had 812 synchronous adenomas and 146 advanced neoplasia. Diminutive, small, and large polyps made up 66.7%, 20.2%, and 13.1% of all lesions, respectively; 9.3% of diminutive and small adenomas were advanced neoplasia, and 45.2% of synchronous advanced lesions were subcentimeter polyps. Both synchronous non-advanced lesions and advanced lesions developed most frequently in the distal colon, followed by the proximal colon, and were least frequently found in the rectum (P < 0.001). Older age (P = 0.04) and male gender (P = 0.001) were associated with the presence of advanced neoplasia in CRC cases with synchronous neoplastic lesions. Meanwhile, the use of aspirin may be associated with a lower incidence of advanced neoplasia (P = 0.04). CONCLUSION: Patients diagnosed with CRC require detailed clearing of the remainder of the colon at baseline coloscopy or postoperative follow-up examination, and we should take a more cautious approach to synchronous subcentimeter polyps in this group of patients.

Detection Rate, Anatomic Sites, and Pathologic Types of Colorectal Cancer During Colonoscopy Procedures.

PURPOSE: This study aims to help physicians obtain the detection rate and colonoscopic information of colorectal cancer (CRC) among patients in a city in China. METHODS: A total of 15,189 participants who underwent total colonoscopy between January 2000 and December 2015 were studied. A total of 1022 CRCs were diagnosed. We analyzed the detection rate, anatomic sites, and pathologic types among different sex, age, and decade groups. Moreover, we investigated the corresponding relationships between the anatomic sites and the pathologic types. RESULTS: Colonoscopic examinations revealed that the risk for CRC between men and women showed no significant difference (6.97% vs. 6.42%). The detection rate of CRC significantly increased with age (2.08% vs. 5.95% vs. 15.09%). The proportion of poorly differentiated adenocarcinoma among the youth group was significantly higher than that in the other age groups (25.0% vs. 11.54% vs. 8.33%). However, the numbers of cases with well-differentiated and moderately differentiated adenocarcinoma observed in the old-aged group were higher than other age groups. This finding demonstrated that the differentiation degree of adenocarcinoma increased with the age. Neuroendocrine tumor was mainly located in the rectum (95.0%), and appeared more frequently among the youth group (7.5% vs. 1.48% vs. 1.06%). CONCLUSIONS: We found the detection rate of CRC varied in terms of sex and decade. The CRC cases in the youth group exhibited a high malignant degree. The most common anatomic site was rectum, so we should focus more on digital rectal examination.

MCM7 expression is correlated with histological subtypes of lung adenocarcinoma and predictive of poor prognosis.

Emerging evidence has implicated that the abnormal expression of MCM3 and MCM7 contributes to tumor formation and progression. However, MCM3 and MCM7 protein expression in different subtypes of lung adenocarcinoma have not yet been reported. In the present study, we detected MCM7 and MCM3 protein level in five subtypes of lung adenocarcinoma by immunohistochemistry. The five subtypes can be divided into 3 grades-grade 1: lepidic adenocarcinoma, grade 2: acinar or papillary adenocarcinoma and grade 3: solid or micropapillary adenocarcinoma. The immunostaining showed that MCM7 level was lowest in the grade 1 subtype and highest in the grade 3 subtypes. The statistical analysis proved that MCM7 expression increased step wisely with the ascending of tumor grades. However, there is no significant relationship between MCM3 expression and tumor grades. In addition, we investigated the association of MCM7 and MCM3 expression with clinicopathological characteristics. The results showed that tumors with lymph node metastasis had higher MCM7 level than those without lymph node metastasis statistically (P<0.0001). MCM3 expression has no significant relationship with clinicopathological characteristics. In conclusion, our results suggested that MCM7 may be a useful biomarker for the pathological diagnosis of subtypes of lung adenocarcinoma and it also may be a potential prognostic marker for lung adenocarcinoma.

MiR-124 inhibits cell proliferation, migration and invasion by directly targeting SOX9 in lung adenocarcinoma.

Accumulating evidence indicates that dysregulation of microRNAs (miRNAs) may contribute to the initiation and progression of cancer. However, the role of miR-124 in lung adenocarcinoma (ADC) and the underlying mechanisms through which miR-124 exerts its functions are not completely understood. In the present study, we detected miR-124 and SOX9 expression in lung ADC tissues. The results showed that miR-124 was significantly downregulated in the lung ADC tissues compared with that noted in the corresponding non-cancerous lung tissues and the level of SOX9 protein was inversely associated with the expression of miR-124. The study in human lung ADC cell line A549 demonstrated that upregulation of miR-124 could inhibit cell proliferation, migration and invasion. The bioinformatic analysis showed that there was a putative miR-124 binding site in the 3' untranslated region (3'UTR) of SOX9. Using a luciferase reporter assay, we verified that SOX9 is a direct target of miR-124. Furthermore, overexpression of miR-124 repressed SOX9 expression, whereas inhibition of miR-124 increased expression of SOX9 in the A549 cells. Finally, we identified that SOX9 was a functional mediator of miR-124 in A549 cells. Taken together, our results suggest that miR-124 functions as a tumor suppressor in lung ADC by directly targeting SOX9 and it may be a promising candidate for miR­based therapy against lung ADC.

Temporal profile of Src, SSeCKS, and angiogenic factors after focal cerebral ischemia: correlations with angiogenesis and cerebral edema.

A better understanding of the underlying mechanisms of angiogenesis and vascular permeability is necessary for the development of therapeutic strategies for ischemic injury. The purpose of this study was to examine the spatial and temporal expression of Src and Src-suppressed C kinase substrate (SSeCKS) in brain after middle cerebral artery occlusion (MCAO) and elucidate the relationships among Src, SSeCKS, and the key angiogenic factors present after stroke. Rats were subjected to either MCAO or sham operation. Reverse transcriptase-polymerase chain reaction and Western blotting results revealed that Src gradually increased starting as early as 2 h after MCAO and remained high for 1 day. In contrast, SSeCKS decreased after MCAO. Src expression correlated positively with that of vascular endothelial growth factor and angiopoietin-2, and negatively with that of SSeCKS, angiopoietin-1, and zonula occludens-1. However, SSeCKS had the reverse correlations. Changes in the expression of these factors correlated with the progress of angiogenesis and cerebral edema. Dynamic temporal changes in Src and SSeCKS expression may modulate angiogenesis and cerebral edema formation after focal cerebral ischemia.