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Owing to their sessile life habit, plants are continuously subjected to a broad range of environmental stresses. During periods of (a)biotic stresses, reactive oxygen species (ROS) levels can rise excessively, leading to oxidative stress. Glutathione reductase (GR) plays an important role in scavenging the ROS and maintenance of redox potential of the cell during oxidative stress. To enhance ROS scavenging capacity, and hence stress tolerance, the Arabidopsis thalianaGR2 (AtGR2) gene was expressed from the tobacco plastid (chloroplast) genome, the main source of ROS production in plant photosynthetic tissues, in this study. Leaves of transplastomic tobacco plants had about seven times GR activity and 1.5 times total glutathione levels compared to wild type. These transplastomic tobacco plants showed no discernible phenotype and exhibited more tolerance to methyl viologen-induced oxidative stress than wild-type control plants. The results indicate that introducing AtGR2 in chloroplasts is an efficient approach to increase stress tolerance. This study also provides evidence that increasing antioxidant enzyme via plastid genome engineering is an alternative to enhance plant’s tolerance to stressful conditions.

Plastid transformation for the expression of recombinant proteins and entire metabolic pathways has become a promising tool for plant biotechnology. However, large-scale application of this technology has been hindered by some technical bottlenecks, including lack of routine transformation protocols for agronomically important crop plants like rice or maize. Currently, there are no standard or commercial plastid transformation vectors available for the scientific community. Construction of a plastid transformation vector usually requires tedious and time-consuming cloning steps. In this study, we describe the adoption of an in vivo Escherichia coli cloning (iVEC) technology to quickly assemble a plastid transformation vector. The method enables simple and seamless build-up of a complete plastid transformation vector from five DNA fragments in a single step. The vector assembled for demonstration purposes contains an enhanced green fluorescent protein (GFP) expression cassette, in which the gfp transgene is driven by the tobacco plastid ribosomal RNA operon promoter fused to the 5′ untranslated region (UTR) from gene10 of bacteriophage T7 and the transcript-stabilizing 3′UTR from the E. coli ribosomal RNA operon rrnB. Successful transformation of the tobacco plastid genome was verified by Southern blot analysis and seed assays. High-level expression of the GFP reporter in the transplastomic plants was visualized by confocal microscopy and Coomassie staining, and GFP accumulation was ~9% of the total soluble protein. The iVEC method represents a simple and efficient approach for construction of plastid transformation vector, and offers great potential for the assembly of increasingly complex vectors for synthetic biology applications in plastids.

BackgroundPathological complete response (pCR) after neoadjuvant chemotherapy (NAC) is associated with favorable outcomes in breast cancer patients. Identifying reliable predictors for pCR can assist in selecting patients who will derive the most benefit from NAC. The prognostic nutritional index (PNI) serves as an indicator of nutritional status and systemic immune competence. It has emerged as a prognostic biomarker in several malignancies; however, its predictive value for pCR in breast cancer remains uncertain. The objective of this study is to assess the predictive value of pretreatment PNI for pCR in breast cancer patients.MethodsA total of 1170 patients who received NAC in two centers were retrospectively analyzed. The patients were divided into three cohorts: a training cohort (n=545), an internal validation cohort (n=233), and an external validation cohort (n=392). Univariate and multivariate analyses were performed to assess the predictive value of PNI and other clinicopathological factors. A stepwise logistic regression model for pCR based on the smallest Akaike information criterion was utilized to develop a nomogram. The C-index, calibration plots and decision curve analysis (DCA) were used to evaluate the discrimination, calibration and clinical value of the model.ResultsPatients with a high PNI (≥53) had a significantly increased pCR rate (OR 2.217, 95% CI 1.215-4.043, p=0.009). Tumor size, clinical nodal status, histological grade, ER, Ki67 and PNI were identified as independent predictors and included in the final model. A nomogram was developed as a graphical representation of the model, which incorporated the PNI and five other factors (AIC=356.13). The nomogram demonstrated satisfactory calibration and discrimination in the training cohort (C-index: 0.816, 95% CI 0.765-0.866), the internal validation cohort (C-index: 0.780, 95% CI 0.697-0.864) and external validation cohort (C-index: 0.714, 95% CI 0.660-0.769). Furthermore, DCA indicated a clinical net benefit from the nomogram.ConclusionThe pretreatment PNI is a reliable predictor for pCR in breast cancer patients. The PNI-based nomogram is a low-cost, noninvasive tool with favorable predictive accuracy for pCR, which can assist in determining individualized treatment strategies for breast cancer patients.

Accurate spinal tuberculosis (TB) diagnosis is of utmost importance for adequately treating and managing the disease. Given the need for additional diagnostic tools, this study aimed to investigate the utility of host serum miRNA biomarkers for diagnosing and distinguishing spinal tuberculosis (STB) from pulmonary tuberculosis (PTB) and other spinal diseases of different origins (SDD). For a case-controlled investigation, a total of 423 subjects were voluntarily recruited, with 157 cases of STB, 83 cases of SDD, 30 cases of active PTB, and 153 cases of healthy controls (CONT) in 4 clinical centers. To discover the STB-specific miRNA biosignature, a high-throughput miRNA profiling study was performed in the pilot study with 12 cases of STB and 8 cases of CONT using the Exiqon miRNA PCR array platform. A bioinformatics study identified that the 3-plasma miRNA combination (hsa-miR-506-3p, hsa-miR-543, hsa-miR-195-5p) might serve as a candidate biomarker for STB. The subsequent training study developed the diagnostic model using multivariate logistic regression in training data sets, including CONT(n=100) and STB (n=100). Youden’s J index determined the optimal classification threshold. Receiver Operating Characteristic (ROC) curve analysis showed that 3-plasma miRNA biomarker signatures have an area under the curve (AUC) = 0.87, sensitivity = 80.5%, and specificity = 80.0%. To explore the possible potential to distinguish spinal TB from PDB and other SDD, the diagnostic model with the same classification threshold was applied to the analysis of the independent validation data set, including CONT(n=45), STB(n=45), brucellosis spondylitis (BS, n=30), PTB (n=30), spinal tumor (ST, n=30) and pyogenic spondylitis (PS, n=23). The results showed diagnostic model based on three miRNA signatures could discriminate the STB from other SDD groups with sensitivity=80%, specificity=96%, Positive Predictive Value (PPV)=84%, Negative Predictive Value (NPV)=94%, the total accuracy rate of 92%. These results indicate that this 3-plasma miRNA biomarker signature could effectively discriminate the STB from other spinal destructive diseases and pulmonary tuberculosis. The present study shows that the diagnostic model based on 3-plasma miRNA biomarker signature (hsa-miR-506-3p, hsa-miR-543, hsa-miR-195-5p) may be used for medical guidance to discriminate the STB from other spinal destructive disease and pulmonary tuberculosis.

BackgroundTo characterize the clinicopathological features and evaluate the neoadjuvant chemotherapy (NACT) efficacy of patients with human epidermal growth factor receptor 2 (HER2)-low breast cancer.MethodsA total of 905 breast cancer patients who received 4 cycles of thrice-weekly standard NACT in the First Affiliated Hospital of Chongqing Medical University were retrospectively enrolled, including 685 cases with HER2-low expression and 220 cases with HER2-negative expression. Clinicopathological features were compared between patients with HER2-negative and HER2-low expression. Univariate and multivariate logistic regression analyses were used to find the independent factors of achieving a pathological complete response (pCR) after NACT.ResultsThere were significant differences in stage_N (P = 0.014), histological grade (P = 0.001), estrogen receptor (ER) status (P < 0.001), progesterone receptor (PgR) status (P < 0.001), NACT regimens (P = 0.032) and NACT efficacy (P = 0.037) between patients with HER2-negative and HER2-low expression breast cancer. In subgroup analysis, histological grade (P = 0.032), ER (P = 0.002), Ki-67 (P < 0.001) and HER2 status (P = 0.025) were independent predictors of achieving a pCR in ER-positive breast cancer. And the nomogram for pCR in ER-positive breast cancer showed great discriminatory ability with an AUC of 0.795. The calibration curve also showed that the predictive ability of the nomogram was a good fit to actual observations. Then, in the analysis of ER-negative breast cancer, only stage_N (P = 0.001) and Ki-67 (P = 0.018) were independent influencing factors of achieving a pCR in ER-negative breast cancer.ConclusionHER2-low breast cancer was a different disease from HER2-negative breast cancer in clinicopathological features. Moreover, the NACT efficacy of HER2-low breast cancer patients was poorer.

PurposeThis study aimed to determine the effect of neoadjuvant chemotherapy (NAC) on circulating levels of reproductive hormones and evaluate the correlation of hormone changes after NAC with hormone receptors expression alterations and relapse-free survival (RFS) outcomes in breast cancer.MethodsInformation from 181 breast cancer patients who received NAC was retrospectively analyzed. For hormones parameters, the median and interquartile range (IQR) were provided at baseline and the end of NAC then was compared by Wilcoxon signed-rank test. Categorical variables were represented as numbers and percentages and were compared via two-sided chi-square and Fisher's tests. The RFS outcomes were compared between patients according to hormone changes using the log-rank test. Univariate and multivariate survival analyses with hazard ratios (HR) and 95% confidence intervals (95% CI) were carried out using Cox regression.ResultsSex steroids including estradiol, progesterone, testosterone, and dehydroepiandrosterone sulfate (DHEAS) levels decreased significantly after NAC among both premenopausal and postmenopausal patients (all P

Ferroptosis, a novel form of regulated cell death (RCD), has garnered increasing attention in studies on numerous human diseases in the last decade. Emerging evidence has indicated that the pathological process of ferroptosis involves the overloaded production of reactive oxygen species (ROS), followed by aberrant accumulation of lipid peroxidation in an iron-dependent manner, accompanied with an increased uptake of polyunsaturated fatty acids into the cellular membrane, further unfolding an ancient vulnerability in multiple context. The unique nature of ferroptosis differentiates it from other forms of RCD, as it is intricately associated with several biological processes, including the metabolism of iron, amino acids, synthesis of ROS and lipid peroxidation. Accordingly, inducers and inhibitors designed to target the key processes of ferroptosis have been extensively studied. Characterized by its distinct properties as mentioned above and its inducible nature, ferroptosis has been widely implicated in several diseases, and numerous studies have focused on identifying effective therapeutic targets for multiple human diseases, including in cancer, by targeting this process. In the present review, recent studies on the involvement of ferroptosis in several types of cancer are summarized and the findings discussed, highlighting the need for increased contemplation of its involvement in the study of cancer, particularly in the clinical setting. A comprehensive summary of the biological mechanisms underlying ferroptosis, the implications of the multiple inducers of ferroptosis, as well as immunotherapy targeting ferroptosis in different types of cancer is provided in this review to highlight the pathophysiological role of ferroptosis in carcinogenesis, to serve as an aid in future studies on the role of ferroptosis in cancer.

BackgroundMetastatic rectal cancer (mRC) of the breast is an extremely rare clinical situation. There are few reported cases in domestic or foreign literature. The clinicopathologic characteristics along with the diagnostic and therapeutic strategies of such cases remain relatively unclear. Here, we would like to provide our comprehensive insights into this rare entity.MethodsWe present a case that till now is the first reported breast metastasis from rectal cancer pathologically diagnosed as a signet-ring cell carcinoma, and we review the current literature on this rare event. The detailed clinical data, histopathology, management, and follow-up aspects were gathered for analysis.ResultsA total of 15 cases were collected including the current case. Breast metastases from rectal cancer present at an average age of 47.7 years (range, 28 to 69 years) and appear with an average interval of 28.4 months (range, 5 months to 18 years) following primary tumor diagnoses. Of the 15 cases, 8 and 5 are pathologically diagnosed as adenocarcinomas and mucinous adenocarcinomas, respectively. Most cases (11/15) are accompanied by extramammary metastases. About half of the breast metastases (7/15) were to the left. In all cases, the main complaints were palpable mass. The average maximum diameter of the metastatic mass is 2.7 cm (range, 1–11 cm). The majority (8/12) of cases with accessible therapy information exclude the option of local surgery.ConclusionPrevious cancer history and accurate immunohistochemistry data play critical roles to distinguish mammary metastasis from a primary neoplasm of the breast. Mastectomy and molecular-targeted drugs should be considered with priority if systemic condition supports them.

Breast cancer (BC) is the second leading cause of death among women and is highly heterogeneous. Three pyroptosis (PR) subtypes were identified in patients with BC from the Cancer Genome Atlas Database (TCGA) cohorts using 20 PR-related regulators, which illustrate a strong association between BC prognosis and PR. Lung metastasis commonly occurs in the advanced stages of BC, resulting in a poor quality of life. Eight differentially expressed (DE) lncRNAs were identified using LASSO–Cox analysis between PR-related and BC lung metastasis. Moreover, a BRCA risk-score (RS) model was established using multivariate Cox regression, which correlated with prognosis in TCGA-BRCA. Clinical characteristics, tumor mutation burden, and tumor immune cell infiltration were extensively investigated between high- and low-RS groups. Similarly, a lower RS implied longer overall survival, greater inflammatory cell infiltration, and better immunotherapeutic response to PD-1 blockers. Our findings provide a foundation for future studies targeting PR and confirme that RS could predict the prognosis of patients with BC.

BackgroundBreast cancer patients who achieve pathological complete response (pCR) after neoadjuvant chemotherapy (NAC) have favorable outcomes. Reliable predictors for pCR help to identify patients who will benefit most from NAC. The pretreatment serum albumin-to-alkaline phosphatase ratio (AAPR) has been shown to be a prognostic predictor in several malignancies, but its predictive value for pCR in breast cancer is still unknown. This study aims to investigate the predictive role of AAPR in breast cancer patients and develop an AAPR-based nomogram for pCR rate prediction.MethodsA total of 780 patients who received anthracycline and taxane-based NAC from January 2012 to March 2018 were retrospectively analyzed. Univariate and multivariate analyses were performed to assess the predictive value of AAPR and other clinicopathological factors. A nomogram was developed and calibrated based on multivariate logistic regression. A validation cohort of 234 patients was utilized to further validate the predictive performance of the model. The C-index, calibration plots and decision curve analysis (DCA) were used to evaluate the discrimination, calibration and clinical value of the model.ResultsPatients with a lower AAPR (

BackgroundHepatocyte growth factor (HGF) signaling plays a plethora of roles in tumorigenesis and progression in many cancer types. As HGF activator inhibitors, serine protease inhibitor, Kunitz types 1 and 2 (SPINT1 and SPINT2) have been reported to be differentially expressed in breast cancer, but their prognostic significance and functioning mechanism remain unclear.MethodsIn our study, multiple databases and bioinformatics tools were used to investigate SPINT1/2 expression profiles, prognostic significance, genetic alteration, methylation, and regulatory network in breast carcinoma.ResultsSPINT1/2 expression was upregulated in breast cancer, and was relatively higher in human epidermal growth factor receptor 2 (HER2) and node positive patients. Elevated SPINT1/2 expression was significantly correlated with a poorer prognosis. Genetic alterations and SPINT1/2 hypomethylation were observed. In breast carcinoma, SPINT1/2 were reciprocally correlated and shared common co-expressed genes. Gene ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis showed that their common co-expressed genes were primarily involved in regulating cell attachment and migration.ConclusionsOur study identified the expression profiles, prognostic significance and potential roles of SPINT1/2 in breast carcinoma. These study results showed that the SPINT1/2 were potential prognostic biomarker for patients with breast cancer.

PurposeThe aim of this study was to assess the prognostic influence of Ki67 index changes in patients with primary triple-negative breast cancer (TNBC) treated with neoadjuvant chemotherapy (NAC), and to evaluate whether the combination of Ki67 index changes and residual disease (RD) tumor-infiltrating lymphocytes (TILs) provides additional prognostic information for this group.Materials and MethodsData from 109 patients with primary TNBC and RD after NAC were analyzed retrospectively. Ki67 changes and RD TIL levels were investigated for associations with recurrence-free survival (RFS) and overall survival (OS) using Kaplan–Meier and Cox analyses.ResultsKi67 index decreased after NAC in 53 patients (48.6%) and high RD TIL levels (≥30%) were observed in 54 patients (49.5%). In multivariate Cox analyses, no Ki67 decrease status and low RD TIL levels were significantly associated with reduced RFS (hazard ratio (HR): 2.038, 95% confidence interval (CI): 1.135–3.658, P = 0.017; HR: 2.493, 95% CI: 1.335–4.653, P = 0.004), and OS (HR: 2.187, 95% CI: 1.173–4.077, P = 0.014; HR: 2.499, 95% CI: 1.285–4.858, P = 0.007), respectively. Notably, low RD TIL levels were significantly associated with reduced RFS (HR: 3.567, 95% CI: 1.475–8.624, P = 0.005) and reduced OS (HR: 3.873, 95% CI: 1.512–9.918, P = 0.005) in only the no Ki67 decrease group. The differences in 3-year RFS and OS between patients with no Ki67 decrease and low or high RD TIL levels were 24.4% vs 79.1% (P = 0.0001) and 33.1% vs 87.5% (P = 0.0001), respectively.ConclusionKi67 index changes and RD TIL levels were associated with the prognosis of patients with primary TNBC with RD after NAC. RD TIL levels had greater prognostic significance in the no Ki67 decrease group.

Breast cancer patients show significant heterogeneity in overall survival. Current assessment models are insufficient to accurately predict patient prognosis, and models for predicting treatment response are lacking. We evaluated the relationship between various immune cells and breast cancer and confirmed the association between immune infiltration and breast cancer progression. Different bioinformatics and statistical approaches were combined to construct a robust immune infiltration-related gene signature for predicting patient prognosis and responses to immunotherapy and chemotherapy. Our research found that a higher immune infiltration-related risk score (IRS) indicates that the patient has a worse prognosis and is not very sensitive to immunotherapy. In addition, a new nomogram was constructed based on the gene signature and clinicopathological features to improve the risk stratification and quantify the risk assessment of individual patients. Our study might contribute to the optimization of the risk stratification for survival and the personalized management of breast cancer.

Background: Triple-negative breast cancer (TNBC) was characterized by breast cancers that do not express estrogen receptor (ER), progesterone receptor (PR), or human epidermal growth factor receptor (HER)-2 genes. TNBC patients are associated with a shorter median time to relapse and death for the lack of available treatment targets. Long non-coding RNAs (LncRNAs) have been reported to play an important role in the development of TNBC. We identified a novel breast-specific long non-coding RNA LINC00993, but less was known about its expression pattern and functional role in TNBC.Methods: LINC00993 RNA expression was detected across different types of clinical breast cancer samples by using qRT-PCR. Bioinformatic methods “guilt by association” and gene set enrichment analysis (GSEA) were used to predict LINC00993 functions. Subcellular localization of LINC00993 in cells was detected by RNA fluorescence in situ hybridization (FISH). Effect of LINC00993 on cell growth was measured by plate colony formation assays, typical growth curve, and an in vivo tumor model. Cell cycle analysis was done by flow cytometry analysis. Key cell cycle regulators were detected by Western blot.Results: LINC00993 was largely downregulated in TNBC, and higher expression indicated better outcome. LINC00993 located mainly in the nucleus. LINC00993 suppressed TNBC growth both in vitro and in vivo. LINC00993 was predicted to be involved in cell cycle pathways by using “guilt by association” and GSEA methods. Key cell cycle regulators like p16INK4A, p14ARF, p53, and p21 were affected by LINC00993 overexpression.Conclusions: A new breast-specific lincRNA LINC00993 was identified with a tumor-suppressive feature and with prognostic value. This is the first research on LINC00993 function. Our results suggest that controlling LINC00993 level may be beneficial for breast cancer treatment.