Branched-Chain Amino Acid Degradation Pathway was Inactivated in Colorectal Cancer: Results from a Proteomics Study.

Background: Colorectal cancer (CRC) ranks third in terms of cancer incidence and fourth in terms of cancer-related deaths worldwide. Identifying potential biomarkers of CRC is crucial for treatment and drug development. Methods: In this study, we established a C57B/6N mouse model of colon carcinogenesis using azoxymethane-dextran sodium sulfate (AOM-DSS) treatment for 14 weeks to identify proteins associated with colon cancer. An isobaric tag for relative and absolute quantitation (iTRAQ)-based proteomic analysis was conducted on the cell membrane components enriched in the colonic mucosa. Additionally, tumor tissues and adjacent normal colon tissues were collected from patients with colon cancer for comparative protein and metabolite analyses. Results: In total, 74 differentially expressed proteins were identified in the tumor tissue samples from AOM/DSS-treated mice compared to both the adjacent tissue samples from AOM/DSS-treated mice and tissue samples from saline-treated control mice. Bioinformatics analysis revealed eight downregulated proteins enriched in the branched-chain amino acids pathway (valine, leucine, and isoleucine degradation). Moreover, these proteins are already known to be associated with the survival rate of patients with cancer. Targeted metabolomics showed increased levels of valine, leucine, and isoleucine in tumor tissues compared to those in adjacent normal tissues in patients with colon cancer. Furthermore, a real-time PCR experiment demonstrated that Aldehyde dehydrogenase, mitochondrial (short protein name ALDH2, gene name Aldh2) and Hydroxyacyl-coenzyme A dehydrogenase, mitochondrial (short protein name HCDH, gene name Hadh) (two genes) in the pathway of branched-chain amino acids) were downregulated in patients with colon cancer (colon tumor tissues vs. their adjacent colon tissues). ALDH2 expression was further validated by western blotting in AOM/DSS-treated mouse model and in clinical samples. Conclusion: This study highlighted the inactivation of the branched-chain amino acid degradation pathway in colon cancer and identified ALDH2 and HCDH as potential biomarkers for diagnosing colon cancer and developing new therapeutic strategies.

Icariin upregulates methyltransferase-like 14-mediated prolyl 4-hydroxylase beta subunit m6A modification to promote osteogenic differentiation of bone marrow stem cells.

Prolyl 4-hydroxylase beta subunit (P4HB) plays a vital role in bone formation. This study intends to clarify the role of P4HB in the therapeutic effect of Icariin (ICA) on osteoporosis. Herein, in vivo and in vitro models were constructed by performing ovariectomy (OVX) in rats and inducing osteogenic differentiation in bone marrow stem cells (BMSCs), respectively. Hematoxylin and eosin staining and micro-computed tomography analysis were performed to evaluate osteoporosis in OVX rats. Alizarin Red staining, alkaline phosphatase staining, and the ALP activity test were employed to assess osteogenesis. m6A dot blotting and methylated RNA immunoprecipitation were used to determine m6A modification. We found that P4HB was downregulated in bone tissues of patients with osteoporosis and OVX rats. P4HB facilitated osteogenic differentiation of BMSCs. What's more, ICA upregulated P4HB expression, promoted osteogenic differentiation of BMSCs, and alleviated osteoporosis in OVX rats, which were reversed by knocking down P4HB. ICA enhanced the stability and m6A modification of P4HB. METTL14 mediated m6A modification of P4HB mRNA. In addition, METTL14 knockdown overturned the promotive effects of ICA on P4HB m6A level and BMSC osteogenic differentiation. To sum up, ICA elevated the METTL14-mediated m6A modification of P4HB to facilitate BMSC osteogenic differentiation.

The value of lactate dehydrogenase to albumin ratio and immune inflammation biomarkers in colorectal cancer.

Background: Colorectal cancer (CRC) is one of the most prevalent gastrointestinal cancers. Evidence for the importance of inflammation and immunology in the development and progression of CRC is growing steadily. The purpose of this study was to determine the clinical importance of Lactic Dehydrogenase (LDH) to Albumin (ALB) Ratio (LAR) and immune-inflammation biomarkers (IIBs) in patients with CRC. Methods: This study enrolled 382 CRC patients. The LAR was determined as the serum LDH(U/l) to ALB(g/l) ratio. We compared the levels of LAR and IIBs in different TNM stages and tumor differentiation. The relationship between LAR and IIBs and overall survival (OS) of CRC was determined by Cox regression models. A prognostic nomogram was created using the results of the multivariate analysis and the effectiveness of the nomogram was assessed using the ROC, calibration, and decision curves. We evaluated the relationship between LAR and IIBs and clinical features of CRC. Results: The levels of LAR, SII, NLR and PLR in TNM IV stage group (LAR:5.92 (5.23-8.24); SII: 1040.02 (499.51-1683.54); NLR: 2.87 (2.07-5.3); PLR:187.08 (125.31-276.63)) were significantly higher than those in other groups. LAR and NLR showed no significant difference in different tumor differentiation groups, while SII and PLR in undifferentiated groups (SII:543.72 (372.63-1110.20); PLR: 147.06 (106.04-203.92)) were significantly higher than those in well and moderate groups (SII: 474.29 (323.75-716.01); PLR: 126.28 (104.31-167.88)). LAR (HR = 1.317, 95% CI = 1.019-1.454), TNM stage (HR = 2.895, 95% CI = 1.838-4.559), age (HR = 1.766, 95% CI = 1.069-2.922) and lymphocytes (HR = 0.663, 95% CI = 0.456-0.963) were predictors of OS. IIBs, including SII, NLR, and PLR are independent of OS. The LAR-based nomogram AUCs of 1-year, 3-year and 5-year survival probabilities in the training cohort were 0.86, 0.72, and 0.71, respectively, and the AUCs of the validation cohort were 0.85, 0.71, and 0.69 respectively. The LAR-based nomogram's ROC curves and calibration curves demonstrated higher OS discriminative performance. The decision curves demonstrated greater net benefit in the survival prediction. Conclusion: Preoperative LAR is a potential prognostic marker in CRC patients, while SII, NLR, and PLR are independent of OS. LAR was associated with tumor stage in CRC patients, but not with tumor differentiation.

PDIA2 Bridges Endoplasmic Reticulum Stress and Metabolic Reprogramming During Malignant Transformation of Chronic Colitis.

Background: Chronic inflammation contributes to approximately 20% of cancers; the underlying mechanisms are still elusive. Here, using an animal model of colitis to colon-cancerous transformation, we demonstrated that endoplasmic reticulum (ER) stress couples with metabolic reprogramming to promote a malignant transformation of chronic inflammation. Methods: The animal model for chronic colitis to colon-cancerous transformation was established in C57BL/6N mice by azoxymethane (AOM) and dextran sodium sulfate (DSS) treatments. The differential proteins in control and AOM/DSS-treated colon mucosa were determined using proteomic analysis; the kinetics of metabolic modifications were monitored by mitochondrial oxygen flux, extracellular acidification, and targeted metabolomics; the molecule linker between ER stress and metabolic modifications were identified by coimmunoprecipitation, KEGG pathway analysis, and the subcutaneous tumor model using gene-specific knockdown colon cancer cells. Tissue array analysis were used to evaluate the differential protein in cancer and cancer-adjacent tissues. Results: AOM/DSS treatment induced 38 tumors in 10 mice at the 14th week with the mean tumor size 9.35 ± 3.87 mm2, which was significantly decreased to 5.85 ± 0.95 mm2 by the ER stress inhibitor 4-phenylbutyric acid (4PBA). Seven differential proteins were determined from control (1,067 ± 48) and AOM/DSS-treated mucosa (1,077 ± 59); the level of ER protein PDIA2 (protein disulfide isomerase-associated 2) was increased over 7-fold in response to AOM/DSS treatment. PDIA2 interacted with 420 proteins that were involved in 8 signaling pathways, in particular with 53 proteins in metabolic pathways. PDIA2 translocated from ER to mitochondria and interacted with the components of complexes I and II to inhibit oxophosphorylation but increase glycolysis. Knockdown PDIA2 in colon cancer cells restored the metabolic imbalance and significantly repressed tumor growth in the xenograft animal model. 4PBA therapy inhibited the AOM/DSS-mediated overexpression of PDIA2 and metabolic modifications and suppressed colon cancer growth. In clinic, PDIA2 was overexpressed in colon cancer tissues rather than cancer-adjacent tissues and was related with the late stages and lymph node metastasis of colon cancer. Conclusions: Persistent ER stress reprograms the metabolism to promote the malignant transformation of chronic colitis; PDIA2 serves as a molecule linker between ER stress and metabolic reprogramming. The inhibition of ER stress restores metabolic homeostasis and attenuates the cancerous transformation of chronic inflammation.

Upregulation of Breast Cancer Resistance Protein Expression was Decreased in Plasma Membrane of Colon Cancer with Metastasis of Lymphatic Node.

BACKGROUND: ATP-binding cassette sub-family G member 2 (ABCG2), an ABC transport protein involved in the efflux of anticancer compounds, has been reported to have altered expression levels in several cancers including breast, colon, and gastric cancer, etc. However, its expression change (up- or down-regulated in cancer) is still contradictory. METHODS: We performed immunohistochemistry to examine ABCG2 expression in the microarray with 90 pairs of colon cancer and their adjacent normal tissues. To find the expression of ABCG2 in lymphatic node metastasis (N1) and N0, we performed immunofluorescence, analyzed by Confocal technology. RESULTS: Compared to adjacent normal tissues, the percentage and density of positive cells expressing ABCG2 in colon cancer were significantly increased. In addition, ABCG2 expression in plasma membrane was related to lymph node metastasis in colon cancer, which was further verified to be downregulated by 2.7-fold in N1 to N0 through immunofluorescent analysis. CONCLUSIONS: ABCG2 expression in colon cancer was up-regulated, and its expression was decreased in the plasma membrane of colon cancer with lymphatic node metastasis (N metastasis).