Characterization of the tumor microenvironment and identification of spatially predictive biomarkers associated with beneficial neoadjuvant chemoradiotherapy in locally advanced rectal cancer.

Neoadjuvant chemoradiotherapy (nCRT) has become the standard treatment for patients with locally advanced rectal cancer (LARC). However, 20-40% of patients with LARC show little to no response to nCRT. Thus, comprehensively understanding the tumor microenvironment (TME), which might influence therapeutic efficacy, and identifying robust predictive biomarkers is urgently needed. Pre-treatment tumor biopsy specimens from patients with LARC were evaluated in detail through digital spatial profiling (DSP), public RNA sequencing datasets, and multiplex immunofluorescence (mIF). DSP analysis revealed distinct characteristics of the tumor stroma compared to the normal stroma and tumor compartments. We identified high levels of human leukocyte antigen-DR/major histocompatibility complex class II (HLA-DR/MHC-II) in the tumor compartment and B cells in the stroma as potential spatial predictors of nCRT efficacy in the Discovery cohort. Public datasets validated their predictive capacity for clinical outcomes. Using mIF in an independent nCRT cohort and/or the total cohort, we validated that a high density of HLA-DR/MHC-II+ cells in the tumor and CD20 + B cells in the stroma was associated with nCRT efficacy (all p ≤ 0.021). Spatial profiling successfully characterized the LARC TME and identified robust biomarkers with the potential to accurately predict nCRT response. These findings have important implications for individualized therapy.

Morroniside alleviates lipopolysaccharide-induced inflammatory and oxidative stress in inflammatory bowel disease by inhibiting NLRP3 and NF-κB signaling pathways.

OBJECTIVE: To investigate the effects of morroniside on inflammatory and oxidative stress in lipopolysaccharide (LPS)-induced inflammatory bowel disease (IBD) cell model. METHODS: NCM460 cells were treated with 2-, 5-, or 10-µg/mL LPS for 24 h to develop an IBD cell model. MTT (3-(4, 5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide) colorimetric assay was performed to uncover the role of morroniside on the viability of LPS-treated NCM460 cells. Flow cytometry and immunoblot assays were performed to confirm the effects of morroniside on the apoptosis of LPS-treated NCM460 cells. Quantitative polymerase chain reaction and enzyme-linked-immunosorbent serologic assays were performed to confirm the effects of morroniside on inflammatory and oxidative stress by measuring the levels of tumor necrosis factor-α, interleukin-1ß, IL-6, superoxide dismutase, malondialdehyde, total antioxidant capacity, and myeloperoxidase. In addition, immunoblot and immunofluorescence assays were performed to detect the effects of morroniside on NLRP3 and NF-κB pathways. RESULTS: Monosine attenuated LPS-induced injury of NCM460 cells. Monosine reduced LPS-induced inflammation in NCM460 cells. In addition, morroniside reduced LPS-induced oxidative stress in NCM460 cells. Mechanically, morroniside suppressed NLRP3 and NF-κB pathways, and alleviated LPS-induced inflammatory and oxidative stress in IBD. CONCLUSION: Morroniside could serve as a promising drug for treating IBD.

A systematic review and quantitative assessment of sleep-disordered breathing during pregnancy and perinatal outcomes.

PURPOSE: Previous investigations have suggested a strong association between sleep-disordered breathing (SDB) during pregnancy and perinatal outcomes. However, the results of the following replication studies were not always concordant. Therefore, this meta-analysis was conducted to evaluate the more reliable estimate. METHODS: A systematic literature search was performed on PubMed, Springer Link, and EMBASE to identify all eligible studies published before August 2013. Summary odds ratios (ORs) and 95% confidence intervals (CIs) were calculated using fixed or random effects model. RESULTS: A total of 24 publications met the inclusion criteria and were included in this meta-analysis. Findings demonstrated that moderate-to-severe SDB during pregnancy was associated with gestational diabetes mellitus (OR=1.78; 95% CI, 1.29 to 2.46), pregnancy-related hypertension (OR=2.38; 95% CI, 1.63 to 3.47), preeclampsia (OR=2.19; 95% CI, 1.71 to 2.80), preterm delivery (OR=1.98; 95% CI, 1.59 to 2.48), low birth weight (OR=1.75; 95% CI, 1.33 to 2.32), neonatal intensive care unit (NICU) admission (OR=2.43; 95% CI, 1.61 to 3.68), intrauterine growth restriction (OR=1.44; 95% CI, 1.22 to 1.71), and Apgar score of <7 at 1 min (OR=1.78; 95% CI, 1.10 to 2.91) based on all studies but not gestational age and birth weight. CONCLUSIONS: This meta-analysis revealed that moderate-to-severe SDB during pregnancy may be associated with most of adverse perinatal outcomes. Further well-designed studies are warranted to confirm our findings.

Heterogeneous expression of mismatch repair proteins and interpretation of immunohistochemical results in colorectal cancer and endometrial cancer.

To investigate the heterogeneous expression patterns of four mismatch repair (MMR) proteins in colorectal cancer (CRC) and endometrial cancer (EC), and their effects on the interpretation of immunohistochemical (IHC) results. A total of 1636 CRC and EC specimens were collected from two hospitals. Seventy-eight cases had heterogeneous expression of MMR proteins, including 49 CRC and 29 EC cases. Polymerase chain reaction-capillary electrophoresis (PCR-CE) was then performed to detect the microsatellite instability (MSI) status, and 44 cases were further verified by targeted next-generation sequencing (NGS). Heterogeneous expression of MMR proteins was observed in 66 cases (66/78, 84.6%) of proficient MMR (pMMR) and 12 cases (12/78, 15.4%) of deficient MMR (dMMR). The proportion of heterogeneous MMR protein expression in EC (12.0%) was higher than that in CRC (3.5%). The heterogeneous expression patterns were divided into focal clonal heterogeneity (6/78, 7.7%) and glandular mosaic heterogeneity (72/78, 92.3%). Surprisingly, three pMMR CRC cases showed isolated small focal clonal heterogeneity of mutS homologue 6 (MSH6), with < 15% positive tumour cells, which was validated as high MSI (MSI-H) with PCR-CE and NGS. However, the other three EC pMMR cases with > 50% focal clonal heterogeneity of MMR proteins were verified as microsatellite stable (MSS) or low MSI (MSI-L). Fifteen EC cases with glandular mosaic heterogeneous expression of MMR proteins included two MSI-H cases, which were validated using PCR-CE and NGS. Among the dMMR cases, only two EC cases with mutL homologue 1 (MLH1)/PMS1 homologue 2, mismatch repair system component (PMS2) loss and MSH2/MSH6 mosaic heterogeneous expression were confirmed as MSS using PCR-CE and NGS, which may be related to the mechanism of MLH1 promoter methylation. Thus, in CRC, only cases with small focal clonal heterogeneous expression of MSH6 have a high likelihood of MSI-H, and further PCR-CE or NGS testing is recommended. The possibility of MSI-H cannot be ruled out in EC cases with glandular mosaic heterogeneous expression of MMR proteins; PCR-CE or NGS detection is therefore necessary.

[Study on effects of microRNA-21 antisense oligonucleotide in vivo and in vitro on bionomics of human cervical squamous carcinoma cell lines SiHa].

OBJECTIVE: To explore the effect of microRNA-21 (miR-21) antisense oligonucleotide on the biological characteristics of human cervical squamous carcinoma cell lines SiHa in vivo and in vitro. METHODS: Specific phosphorothioate antisense oligodeoxynucleotides targeting miR-21 were synthesized and transfected into cervical cancer cells in vitro. Expression of miR-21 in SiHa after transfection was detected by real-time RT-PCR. The cell proliferation was evaluated by MTT assay and colony formation experiment. The cell apoptosis was analyzed by annexin V-FITC/PI analysis. The inhibitory effect of miR-21 antisense oligonucleotide on tumor growth was evaluated by tumor growth curves and immunohistochemistry (MaxVision method). H-E staining was used to document morphological changes and fluorometric TUNEL assay was to detect the apoptotic activity. RESULTS: After the transfection of antisense miR-21, the expression of miR-21 decreased along with an obvious growth inhibition, compared with that of the control groups (P < 0.05). Colony formation of both cell lines was markedly inhibited with antisense miR-21 (55.6% ± 1.4%), as compared with that in the negative group (98.3% ± 2.0%, P < 0.05). Flow cytometry assay showed that antisense miR-21 expression significantly enhanced the cell apoptosis (6.7% ± 1.3% and 29.4% ± 1.7%, P < 0.05). The tumor-forming rates of miR-21 transfected group, and negative control groups were 3/8 and 6/8, respectively (P < 0.05). Ki-67 proliferative marker staining decreased significantly (42% vs 90%) in the transfected group compared with negative control groups. Extensive dead tumor cells were seen in the miR-21 transfected cells along with a marked increase of apoptosis (P < 0.05). CONCLUSION: Targeted antisense oligonucleotide miR-21 effectively suppresses the growth of cervical carcinoma SiHa cells both in vitro and in vivo through an induction of apoptosis.

[MicroRNA383 regulates expression of PRDX3 in human medulloblastomas].

OBJECTIVE: To investigate the effects of microRNA-383 (miR-383) on PRDX3 gene expression, cell proliferation and apoptosis of human medulloblastma. METHODS: PRDX3 and miR-383 RNA expression was detected by real-time quantitative RT-PCR in human medulloblastoma tumor tissue samples, Daoy cell line and normal brain tissue samples. Western blot was used to detect protein expression of PRDX3. Synthetic miR-383 mimics were transfected into Daoy cells by lipofectamine. Using Cell Counting Kit-8 (CCK-8) method, flow cytometry was used to investigate the cell proliferation and apoptosis, cells reactive oxgen species(ROS), mitochondrial membrane potential changes in each experimental groups. RESULTS: Of 15 cases of human medulloblastoma tumor, 13 cases had miR-383 expression levels significantly lower than that of normal brain tissue, and 14 had PRDX3 mRNA expression levels significantly higher than that of normal brain tissue. The expression levels of miR-383 and PRDX3 in Daoy cells were 0.353 and 1.315 times than those of normal brain tissue, respectively. The protein expression levels of PRDX3 were higher in human medulloblatoma tumors and Daoy cells than that of normal brain tissue. Transfected miR-383 mimics increased the expression level of miR-383 after 24 h and 48 h was significantly higher than that of the control. In contrast, PRDX3 gene mRNA and protein expression levels were significantly decreased at 48 h compared with the control group. Using CCK-8 assay, the cell proliferation rate in the experimental group was significantly lower than that of the control group (P < 0.05). Annexin V-FITC assay demonstrated that early apoptosis rate of the experimental group (11.60 ± 0.30)% was significantly higher than those of the control group (2.3 ± 0.20)% and negative control group (10.37 ± 0.25)% (P = 0.000) after 48 h of transfection. The intracellular ROS levels after transfection at 24 and 48 h significantly increased than those of the control group. Mitochondrial membrane potential level at 24 h after transfection significantly decreased, comparing with the blank control group and the negative control group. CONCLUSIONS: Compared with normal brain tissue, decreased expression of miR-383 but elevated expression of PRDX3 are medulloblastoma tumour and Daoy cell lines. Up-regulation of miR-383 knockdowns the expression of PRDX3, inhibits proliferation and promotes apoptosis of Daoy cells, leading to increased intracellular ROS and decreased levels of mitochondrial membrane potential.

MicroRNA­30a regulates cell proliferation, migration, invasion and apoptosis in human nasopharyngeal carcinoma via targeted regulation of ZEB2.

MicroRNA­30a (miR­30a) was previously reported to serve as a tumor suppressor able to inhibit the development and progression of certain types of cancer. A number of previous studies demonstrated that zinc finger E­box binding homeobox 2 (ZEB2) may be regulated by miR­30a in clear cell renal cell carcinoma and breast cancer. However, the function of miR­30a in human nasopharyngeal carcinoma (NPC) remains unclear. The present study aimed to investigate the association between miR­30a and ZEB2 in NPC. Therefore, the expression levels of miR­30a and ZEB2 were measured in human NPC cells and tissues from patients with NPC, and the present results suggested that the expression level of miR­30a was significantly decreased in NPC tissues compared with paracancerous tissues. The direct interaction between miR­30a and the untranslated region of ZEB2 was examined using the dual­luciferase reporter assay, and ZEB2 was identified as a direct target of miR­30a. Additionally, the effects of miR­30a and ZEB2 overexpression on cell proliferation, migration, invasion and apoptosis were additionally investigated. Functional experiments identified that overexpression of miR­30a increased apoptosis and suppressed cell proliferation, cell migration and cell invasion by directly targeting ZEB2. Collectively, the present study suggested that miR­30a may serve an important role in the progression of NPC and may represent a novel target for the treatment of patients with NPC.

Maternal anxiety during pregnancy and adverse birth outcomes: a systematic review and meta-analysis of prospective cohort studies.

BACKGROUND: Previous studies concerning the association between maternal anxiety during pregnancy and adverse birth outcomes have provided controversial findings. METHODS: In this systematic review, a meta-analysis was utilized to investigate the association between maternal anxiety and preterm birth (PTB) and/or low birth weight (LBW). Literature was searched until June 2013. Only prospective cohort studies that reported data on maternal anxiety during pregnancy with PTB and/or LBW were included. Pooled relative risks (RRs) with 95% confidence intervals (CIs) were calculated using fixed or random effects models depending on the size of heterogeneity. RESULTS: Twelve studies totaling 17,304 pregnant women reported PTB data; and six studies totaling 4948 pregnant women reported LBW data. Maternal anxiety during pregnancy was associated with significant increased risk of PTB (pooled RR=1.50, 95% CI=1.33-1.70) and LBW (pooled RR=1.76, 95% CI=1.32-2.33). LIMITATIONS: Potential moderators could not be adequately considered due to insufficient information. In addition, the effects of different types of anxiety disorder on the risk of these adverse birth outcomes could not be investigated. CONCLUSIONS: The results suggested that maternal anxiety during pregnancy was positively related to an increased risk of PTB and LBW. Healthcare providers should give close attention to anxiety in pregnant women and provide appropriate mental health support in order to improve outcomes for both mothers and infants.