Methylated ctDNA predicts early recurrence risk in patients undergoing resection of initially unresectable colorectal cancer liver metastases.

Background: Patients with initially unresectable colorectal cancer liver metastases (IU-CRLM) might benefit from using an effective systemic treatment followed by resection of liver metastases but the curative success rate is quite low. Indeed, nearly one-third of patients exhibit early recurrence within the first 6 months after surgery, and these individuals often have poor overall survival. Objectives: This study aims to clarify the application value of serial circulating tumor DNA (ctDNA) analysis in predicting the clinical outcome of IU-CRLM patients following liver metastasectomy. Design: A retrospective study was conducted on a cohort of patients with IU-CRLM between February 2018 and April 2021. Methods: Plasma samples at different time points during CRLM treatment [baseline (BL), preoperation (PRE), postoperation (POST), end-of-treatment (EOT), and progressive disease (PD)] were retrospectively collected from patients with initially unresectable CRLM enrolled at the Sun Yat-sen University Cancer Center. Dynamic changes of SEPTIN 9 (SEPT9) and Neuropeptide Y (NPY) methylated circulating tumor DNA (MetctDNA) levels in serial plasma samples were detected using droplet-digital PCR (ddPCR). Results: SEPT9 and NPY genes were hypermethylated in colon cancer cell lines and tissues while no difference was observed between primary and metastatic tumors. Patients with MetctDNA positive at POST or EOT had significantly lower recurrence-free survival (RFS) compared to patients with MetctDNA negative at these time points [POST: Hazard ratio (HR) 9.44, 95% confidence interval (CI) 5.15-17.30, p < 0.001; EOT: HR 11.48, 95% CI 3.27-40.31, p < 0.001]. Multivariate analysis demonstrated that POST (OR 33.96, 95% CI 4.03-286.10, p = 0.001) and EOT (OR 18.36, 95% CI 1.14-295.71, p = 0.04) MetctDNA was an independent risk factor for early recurrence. Time-dependent receiver operating characteristic curve (T-ROC) analysis revealed that area under the curve (AUC) value was greatest at the relapse time point of 6 months post-intervention, with POST-AUC and EOT-AUC values of 0.74 (95% CI 0.66-0.81) and 0.73 (95% CI 0.53-0.94), respectively. Serial MetctDNA analysis showed that RFS was significantly lower in patients with no MetctDNA clearance compared with those with MetctDNA clearance (HR 26.05, 95% CI 4.92-137.81, p < 0.001). Conclusion: Our study confirmed that serial ctDNA analysis of NPY and SEPT9 gene methylation could effectively predict early recurrence in IU-CRLM patients, especially at POST and EOT.

Changes in inflammatory markers and efficacy analysis of continuous subcutaneous insulin infusion in senior patients with type 2 diabetes mellitus hospitalized with community-acquired pneumonia: a randomized controlled trial.

OBJECTIVE: The purpose of this study is to analyze the changes in inflammatory markers and efficacy in the treatment of senior patients with type 2 diabetes mellitus (T2DM) and community-acquired pneumonia with continuous subcutaneous insulin infusion (CSII). METHODS: A total of 105 senior patients with T2DM and community-acquired pneumonia, were randomly divided into two groups, viz., treatment group and control group-52 patients in the treatment group were treated with CSII, and 53 patients in the control group with multiple daily insulin injections (MDI). The changes in fasting blood glucose, postprandial blood glucose, total number of white blood cells, neutrophils, percentage of neutrophils, lymphocytes, percentage of lymphocytes, C-reactive protein, serum amyloid A, procalcitonin, interleukin-6 indexes, and the improvement in clinical outcome between the two groups were compared on the 5th and the 10th day of treatment. RESULTS: In the treatment group, there were 52 patients with an average age of (73.7 ± 8.5) years, which included 28 males and 24 females. In the control group, there were 53 patients, with 27 males and 26 females, with an average age of (74.8 ± 8.8) years. On the 5th and the 10th day of the treatment, the fasting blood glucose, postprandial blood glucose, total number of white blood cells, neutrophils, percentage of neutrophils, lymphocytes, percentage of lymphocytes, C-reactive protein, serum amyloid A, procalcitonin and interleukin-6 of the treatment group were better than that of the control group (P < 0.05). The use of CSII was associated with a higher probability of a prompt recovery (P < 0.05). CONCLUSION: The administration of CSII in the treatment of senior patients with T2DM and community-acquired pneumonia can effectively control fasting and postprandial blood glucose, significantly reduce the levels of inflammatory markers, and improve infection treatment efficacy.

Response to programmed cell death protein 1 antibody in patients with Epstein-Barr virus-associated intrahepatic cholangiocarcinoma.

AIM: Epstein-Barr virus-associated intrahepatic cholangiocarcinoma (EBVaICC) has a distinct genomic profile and increased CD3+ and CD8+ T cells infiltration. However, the efficacy of immunotherapy in EBVaICC remains largely unknown. This study aimed to assess the efficacy of programmed cell death protein 1 (PD-1) antibody therapy in EBVaICC. METHODS: Patients with metastatic biliary tract cancer (BTC) diagnosed at Sun Yat-sen University Cancer Center from January 2016 to December 2021 were identified. In situ hybridisation was performed to detect EBV. Overall survival (OS) and progression-free survival (PFS) were measured. RESULTS: A total of 698 patients with metastatic BTC were identified, of whom 39 (5.6%) had EBVaICC. Among the 136 patients who were not administered PD-1 antibody, the OS was similar between patients with EBVaICC and EBV-negative ICC (median OS 12.5 versus 9.5 months, respectively; P = 0.692). For the 205 patients who were administered PD-1 antibody, patients with EBVaICC had significantly longer OS than patients with EBV-negative ICC (median OS 24.9 versus 11.9 months, respectively; P = 0.004). Seventeen patients with EBVaICC were administered PD-1 antibody. Eight patients (47%) achieved a partial response, and 17 patients achieved disease control. The median PFS was 17.5 months. CONCLUSIONS: This study identified a clinically actionable subset of patients with EBVaICC with a promising response to the PD-1 antibody.

Adipose-derived mesenchymal stem cell exosomes ameliorate spinal cord injury in rats by activating the Nrf2/HO-1 pathway and regulating microglial polarization.

As of now, there are no satisfactory treatments for spinal cord injury (SCI), so new therapeutic approaches are necessary to be explored. Adipose-derived mesenchymal stem cell exosomes (ADMSC-Exo), delightfully, show remarkable therapeutic effects. Therefore, we try to investigate the effects and mechanisms of ADMSC-Exo on SCI, as well as to provide novel approaches for the treatment of SCI. Adipose-derived mesenchymal stem cells (ADMSC) were isolated from rats and then exosomes (Exo) were extracted from the cells. The extracted Exo were identified by flow cytometry, transmission electron microscopy and nanoparticle tracking analysis (NTA). Then, the SCI rat model was established by the spinal cord impactor and injected with 200 µl PBS or Exo into their tail veins at 30 min, 24 h, and 48 h after surgery. The rats in the Control group and Exo group only exposed the spine. Motor function recovery was assessed on days 0, 7, 14, 21, and 28; histopathological changes and apoptosis levels in spinal cord tissues were observed by HE staining and TUNEL staining; the levels of inflammatory cytokines TNF-a, IL-6, and MCP-1 in spinal cord tissues were measured by ELISA; the expression levels of iNOS, IL-12, Arg1, and Mrc1 in spinal cord tissues were detected by qRT-PCR; and Nrf2, HO-1, and NQO1 protein expression in spinal cord tissues were detected by Western blot. ADMSC-Exo were successfully isolated and identified. ADMSC-Exo significantly relieved SCI and promoted motor function recovery in SCI rats. Moreover, ADMSC-Exo inhibited the expression of both inflammatory factors in the spinal cord tissues and M1 microglia, promoted the expression of M2 microglia, and activated the Nrf2/HO-1 pathway. Altogether, ADMSC-Exo can not only ameliorate SCI, but also promote the motor function recovery of rats. And the mechanism of ADMSC-Exo improving SCI may be achieved by activating Nrf2/HO-1 pathway and microglial polarization.

Targeted demethylation of the CDO1 promoter based on CRISPR system inhibits the malignant potential of breast cancer cells.

BACKGROUND: Cysteine dioxygenase 1 (CDO1) is frequently methylated, and its expression is decreased in many human cancers including breast cancer (BC). However, the functional and mechanistic aspects of CDO1 inactivation in BC are poorly understood, and the diagnostic significance of serum CDO1 methylation remains unclear. METHODS: We performed bioinformatics analysis of publicly available databases and employed MassARRAY EpiTYPER methylation sequencing technology to identify differentially methylated sites in the CDO1 promoter of BC tissues compared to normal adjacent tissues (NATs). Subsequently, we developed a MethyLight assay using specific primers and probes for these CpG sites to detect the percentage of methylated reference (PMR) of the CDO1 promoter. Furthermore, both LentiCRISPR/dCas9-Tet1CD-based CDO1-targeted demethylation system and CDO1 overexpression strategy were utilized to detect the function and underlying mechanism of CDO1 in BC. Finally, the early diagnostic value of CDO1 as a methylation biomarker in BC serum was evaluated. RESULTS: CDO1 promoter was hypermethylated in BC tissues, which was related to poor prognosis (p < .05). The CRISPR/dCas9-based targeted demethylation system significantly reduced the PMR of CDO1 promotor and increased CDO1 expression in BC cells. Consequently, this leads to suppression of cell proliferation, migration and invasion. Additionally, we found that CDO1 exerted a tumour suppressor effect by inhibiting the cell cycle, promoting cell apoptosis and ferroptosis. Furthermore, we employed the MethyLight to detect CDO1 PMR in BC serum, and we discovered that serum CDO1 methylation was an effective non-invasive biomarker for early diagnosis of BC. CONCLUSIONS: CDO1 is hypermethylated and acts as a tumour suppressor gene in BC. Epigenetic editing of abnormal CDO1 methylation could have a crucial role in the clinical treatment and prognosis of BC. Additionally, serum CDO1 methylation holds promise as a valuable biomarker for the early diagnosis and management of BC.

Phosphoserine phosphatase as a prognostic biomarker in patients with gastric cancer and its potential association with immune cells.

BACKGROUND: Because of dismal prognosis in gastric cancer, identifying relevant prognostic factors is necessary. Phosphoserine phosphatase (PSPH) exhibits different expression patterns in many cancers and has been reported to affect the prognosis of patients with cancer. In this study, we examined the prognostic role of metabolic gene PSPH in gastric cancer based on the TCGA dataset and our hospital-based cohort cases. METHODS: We collected and analysed RNA-seq data of Pan-cancer and gastric cancer in the TCGA dataset and PSPH expression data obtained from immunohistochemical analysis of 243 patients with gastric cancer from Sun Yat-sen University cancer center. Further, Kaplan-Meier survival analysis and Cox analysis were used to assess the effect of PSPH on prognosis. The ESTIMATE and Cibersort algorithms were used to elucidate the relationship between PSPH and the abundance of immune cells using the TCGA dataset. RESULTS: We observed that PSPH expression displayed considerably high in gastric cancer and it was significantly associated with inferior prognosis (P = 0.043). Surprisingly, there was a significant relationship between lower immune scores and high expression of PSPH (P < 0.05). Furthermore, patients with a low amount of immune cells exhibited poor prognosis (P = 0.046). The expression of PSPH significantly increased in activated memory CD4 T cells, resting NK cells and M0 macrophages (P = 0.037, < 0.001, and 0.005, respectively). CONCLUSIONS: This study highlighted that PSPH influences the prognosis of patients with gastric cancer, and this is associated with the infiltration of tumour immune cells, indicating that PSPH may be a new immune-related target for treating gastric cancer.

Mutational landscape of primary pulmonary salivary gland-type tumors through targeted next-generation sequencing.

OBJECTIVES: Primary pulmonary salivary gland-type tumors (PSGTs) mainly comprise of mucoepidermoid carcinoma (MEC) and adenoid cystic carcinoma (ACC), which are rare and molecularly poorly understood. This study aimed to profile the molecular alterations of PSGTs via targeted next-generation sequencing (NGS). MATERIAL AND METHODS: Immunohistochemistry was used to screen PSGTs in 32 patients and MAML2 and MYB rearrangements were detected using fluorescence in situ hybridization. 1021-Genepanel of targeted NGS was conducted to profile genomic mutations in all the PSGT patients. RESULTS: Among the 32 patients, 25 had MEC and 7 had ACC. MAML2 and MYB rearrangements were detected in 80.0% (20/25) of the MEC and 71.4% (5/7) of the ACC patients. Among the MEC patients, 10 (40.0%) had ≥1 mutation, and 6 of them had 11 isolated mutations with abundance >5%, namely NFE2L2, MYOD1, INPP4B, CCND2, SNTG1, HSPD1, TGFBR1, RBM10, NOTCH4, ASXL1, and PTPRD mutations. The remaining 4 patients had 9 mutations with abundance <5%, namely KMT2A, PDCD11, FLT1, BRCA2, APC, SLX4, FOXP1, FGFR1, and HRAS mutations. All the ACC patients had mutations, which were enriched in 5 pathways including the PI3K and NOTCH pathways, chromatin and cytoskeleton remodeling, and DNA damage. These results explain PSGTs harbor distinct driver features of MAML2 or MYB rearrangement, accompanied with wide mutational diversity with very low rate of somatic mutation. Several important pathways, including the NOTCH and PI3K pathways, and chromatin remodeling could be targeted to improve the survival in patients with ACC.

Plasma SGIP1 methylation in diagnosis and prognosis prediction in hepatocellular carcinoma.

Aberrant methylation of some genes can serve as promising biomarkers in hepatocellular carcinoma (HCC). This study aimed to investigate the diagnostic and prognostic value of plasma SGIP1 methylation in HCC. The study included a total of 269 subjects, of which 129 were with HCC, 45 with liver cirrhosis (LC), 45 with chronic hepatitis B (CHB), and 50 were healthy controls (HCs). The aberrant methylation was detected by quantitative methylation-specific polymerase chain reaction (qMSP). The area under the curve (AUC) was 0.872 in distinguishing HCC from HCs, with a sensitivity of 85.3% and a specificity of 88%. The AUC was 0.728, when it distinguished HCC from CHB, with a sensitivity of 43.4% and a specificity of 97.8%. The AUC was 0.728 in distinguishing HCC from LC, with a sensitivity of 43.4% and a specificity of 97.8%. Elevated levels of SGIP1 methylation in HCC patients showed poorer overall survival (OS), progression-free survival (PFS), and metastasis-free survival (MFS) than those with low levels (Kaplan-Meier method and the log-rank test, p<0.05). SGIP1 methylation in different study groups demonstrated different sensitivities. SGIP1 methylation detection in the plasma may serve as a non-invasive diagnostic and prognostic biomarker for HCC.

Association between XRCC1 polymorphisms and glioma risk among Chinese population.

The pathogenesis of glioma remains largely unknown now. It has been suggested that the X-ray cross-complementing group 1 (XRCC1) gene may influence the capacity to repair DNA damage leading to an increased gliomas susceptibility. This study aimed to evaluate the relationship between XRCC1 polymorphisms and glioma risk. Genotypes were assessed in 368 Chinese glioma patients and 346 healthy controls. XRCC1 Arg194Trp (rs1799782), Arg280His (rs25489) and Arg399Gln (rs25487) and three additional polymorphisms were directly sequenced. The frequency of Arg280His A allele was significant lower in glioma group than in healthy controls [9.6 vs 16%, OR=0.60 (0.46-0.80), P<0.001]; the frequencies of GA or AA genotypes were different in two groups (16.6 vs 22.8%, 1.3 vs 4.7%). The frequency of Arg399Gln A allele was significant higher in glioma group than in healthy controls [38.7 vs 30.1%, OR=1.29 (1.11-1.49), P=0.001]; the frequencies of GA or AA genotypes were different in two groups (45.4 vs 38.2%, 16 vs 10.9%). This study demonstrates that the rs25489 (Arg280His) and Arg399Gln (rs25487) polymorphisms in XRCC1 gene might influence the risk of developing glioma in Chinese population.

[Localization and expression pattern of MDM2 in axon initial segments of neuron in rodent brain].

To investigate the murine double minute 2 (MDM2) localization and expression pattern in brain, immunohistochemistry, immunofluorescent staining and immunoblotting methods were used to analyze it in brains of Kunming mice during postnatal development, in brains of adult SD rats and in primarily cultured neurons. The distribution of MDM2 and markers of axon initial segment (AIS) was analyzed by double immunolabeling. In addition, Nutlin-3, a MDM2 antagonist, was injected into hippocampus to analyze the effect on the distribution of MDM2 and AIS protein Nav1.6 in AIS. The results showed that the dynamic expression patterns of MDM2 protein in cerebral cortex and hippocampus of Kunming mice after birth were different. However, it was similar that MDM2 was gradually enriched to AIS during postnatal development, especially after postnatal day 7. The MDM2 in AIS was also observed in different brain regions of adult SD rat brain and in primarily cultured neurons, where MDM2 was colocalized with AIS markers such as AnkG and Nav1.6. In addition, hippocampal injection of Nutlin-3 could induce the loss of the characteristic distribution of MDM2 in AIS. Moreover, Nutlin-3 not only caused a decrease of Nav1.6 distributing in AIS, but also disrupted the polarized distribution of MAP2 in neurons. These results indicate that MDM2 can be enriched at the AIS of adult rodent brain, which might play a role in regulation of the maintenance of AIS function and neuronal polarity.

Upregulation of heme oxygenase-1 by acteoside through ERK and PI3 K/Akt pathway confer neuroprotection against beta-amyloid-induced neurotoxicity.

Our previous study has shown that acteoside, an antioxidative phenylethanoid glycoside, protect against beta-amyloid (Aß)-induced cytotoxicity in vitro. However, the precise protective mechanisms remains unclear. Heme oxygenase-1 (HO-1) is a crucial factor in the response to oxidative injury, protecting neurons against Aß-induced injury. In the present study we examined to determine whether acteoside upregulates HO-1 expression, and thereby protects PC12 cells against Aß-induced cell death. It was revealed that acteoside is an activator of Nrf2 and inducer of HO-1 expression. We showed that acteoside increased HO-1 expression in vitro and in vivo. Acteoside treatment resulted in nuclear translocation of the transcription factor NF-E2-related factor 2 (Nrf2). Acteoside activated both ERK and PI3 K/Akt, and treatments with the specific ERK inhibitor PD98059, the PI3 K inhibitor LY294002, and the specific Nrf2 siRNA suppressed the acteoside-induced HO-1 expression. The HO-1 inhibitor ZnPP, PD98059, and LY294002 markedly abolished the neuroprotective effect of acteoside against Aß-induced neurotoxicity. Taken together, these results demonstrate that acteoside is an activator of Nrf2 and inducer of HO-1 expression. We also showed that acteoside increased HO-1 expression through activation of ERK and PI3 K/Akt signal pathways in vitro. Upregulation of HO-1 by acteoside may involve in the neuroprotection against Aß-induced neurotoxicity.

Overexpression of F(0)F(1)-ATP synthase alpha suppresses mutant huntingtin aggregation and toxicity in vitro.

Huntington's disease (HD) and other polyglutamine (polyQ) neurodegenerative diseases are characterized by neuronal accumulation of the disease protein, suggesting that the cellular ability to handle abnormal proteins is compromised. As a multi-subunit protein localized in the mitochondria of eukaryotic cells, the F(0)F(1)-ATP synthase alpha belongs to the family of stress proteins HSP60. Currently, mounting evidences indicate F(0)F(1)-ATP synthase alpha may play a role in neurodegenerative diseases, including Alzheimer's disease (AD) and Parkinson's disease (PD). Recently, ATP synthase alpha was reported to have protective and therapeutic roles in primary cardiacmyocytes of iron-overloaded rats by lowering ROS production. However, little is understood about the role of ATP synthase alpha in cell death and neurodegeneration. Here, we demonstrate that overexpression of ATP synthase alpha suppresses huntingtin (htt) polyQ aggregation and toxicity in transfected SH-SY5Y cell lines. Overexpression of ATP synthase alpha is able to protect cell death caused by polyglutamine-expanded htt. Transient overexpression of ATP synthase alpha suppresses the aggregate formation by estimation of polyQ aggregation, Western blot analysis, and filter trap assay (FTA) in transfected SH-SY5Y cells. These results indicated that ATP synthase alpha has a strong inhibitory effect on polyglutamine aggregate formation and toxicity in vitro, and suggest a novel neuroprotective role of ATP synthase alpha.