Integrated whole-exome and bulk transcriptome sequencing delineates the dynamic evolution from preneoplasia to invasive lung adenocarcinoma featured with ground-glass nodules.

OBJECTIVE: The genomic and molecular ecology involved in the stepwise continuum progression of lung adenocarcinoma (LUAD) from adenocarcinoma in situ (AIS) to minimally invasive adenocarcinoma (MIA) and subsequent invasive adenocarcinoma (IAC) remains unclear and requires further elucidation. We aimed to characterize gene mutations and expression landscapes, and explore the association between differentially expressed genes (DEGs) and significantly mutated genes (SMGs) during the dynamic evolution from AIS to IAC. METHODS: Thirty-five patients with ground-glass nodules (GGNs) lung adenocarcinomas were enrolled. Whole-exome sequencing (WES) and transcriptome sequencing (RNA-Seq) were conducted on all patients, encompassing both tumor samples and corresponding noncancerous tissues. Data obtained from WES and RNA-Seq were subsequently analyzed. RESULTS: The findings from WES delineated that the predominant mutations were observed in EGFR (49%) and ANKRD36C (17%). SMGs, including EGFR and RBM10, were associated with the dynamic evolution from AIS to IAC. Meanwhile, DEGs, including GPR143, CCR9, ADAMTS16, and others were associated with the entire process of invasive LUAD. We found that the signaling pathways related to cell migration and invasion were upregulated, and the signaling pathways of angiogenesis were downregulated across the pathological stages. Furthermore, we found that the messenger RNA (mRNA) levels of FAM83A, MAL2, DEPTOR, and others were significantly correlated with CNVs. Gene set enrichment analysis (GSEA) showed that heme metabolism and cholesterol homeostasis pathways were significantly upregulated in patients with EGFR/RBM10 co-mutations, and these patients may have poorer overall survival than those with EGFR mutations. Based on the six calculation methods for the immune infiltration score, NK/CD8+ T cells decreased, and Treg/B cells increased with the progression of early LUAD. CONCLUSIONS: Our findings offer valuable insights into the unique genomic and molecular features of LUAD, facilitating the identification and advancement of precision medicine strategies targeting the invasive progression of LUAD from AIS to IAC.

Lactate-upregulated NADPH-dependent NOX4 expression via HCAR1/PI3K pathway contributes to ROS-induced osteoarthritis chondrocyte damage.

Increasing evidence shows that metabolic factors are involved in the pathological process of osteoarthritis (OA). Lactate has been shown to contribute to the onset and progression of diseases. While whether lactate is involved in the pathogenesis of OA through impaired chondrocyte function and its mechanism remains unclear. This study confirmed that serum lactate levels were elevated in OA patients compared to healthy controls and were positively correlated with synovial fluid lactate levels, which were also correlated with fasting blood glucose, high-density lipoprotein, triglyceride. Lactate treatment could up-regulate expressions of the lactate receptor hydroxy-carboxylic acid receptor 1 (HCAR1) and lactate transporters in human chondrocytes. We demonstrated the dual role of lactate, which as a metabolite increased NADPH levels by shunting glucose metabolism to the pentose phosphate pathway, and as a signaling molecule up-regulated NADPH oxidase 4 (NOX4) via activating PI3K/Akt signaling pathway through receptor HCAR1. Particularly, lactate could promote reactive oxygen species (ROS) generation and chondrocyte damage, which was attenuated by pre-treatment with the NOX4 inhibitor GLX351322. We also confirmed that lactate could increase expression of catabolic enzymes (MMP-3/13, ADAMTS-4), reduce the synthesis of type II collagen, promote expression of inflammatory cytokines (IL-6, CCL-3/4), and induce cellular hypertrophy and aging in chondrocytes. Subsequently, we showed that chondrocyte damage mediated by lactate could be reversed by pre-treatment with N-Acetyl-l-cysteine (NAC, ROS scavenger). Finally, we further verified in vivo that intra-articular injection of lactate in Sprague Dawley (SD) rat models could damage cartilage and exacerbate the progression of OA models that could be countered by the NOX4 inhibitor GLX351322. Our study highlights the involvement of lactate as a metabolic factor in the OA process, providing a theoretical basis for potential metabolic therapies of OA in the future.

Early-onset locally advanced rectal cancer characteristics, a practical nomogram and risk stratification system: a population-based study.

Background: The purpose of this study is to construct a novel and practical nomogram and risk stratification system to accurately predict cancer-specific survival (CSS) of early-onset locally advanced rectal cancer (EO-LARC) patients. Methods: A total of 2440 patients diagnosed with EO-LARC between 2010 and 2019 were screened from the Surveillance, Epidemiology, and End Results (SEER) database. The pool of potentially eligible patients was randomly divided into two groups: a training cohort (N=1708) and a validation cohort (N=732). The nomogram was developed and calibrated using various methods, including the coherence index (C-index), receiver operating characteristic curve (ROC), calibration curves, and decision curves (DCA). A new risk classification system was established based on the nomogram. To compare the performance of this nomogram to that of the American Joint Committee on Cancer (AJCC) staging system, DCA, net reclassification index (NRI), and integrated discrimination improvement (IDI) were employed. Result: Seven variables were included in the model. The area under the ROC curve (AUC) for the training cohort was 0.766, 0.736, and 0.731 at 3, 6, and 9 years, respectively. Calibration plots displayed good consistency between actual observations and the nomogram's predictions. The DCA curve further demonstrated the validity of the nomination form in clinical practice. Based on the scores of the nomogram, all patients were divided into a low-risk group, a middle-risk group, and a high-risk group. NRI for the 3-, 6-, and 9-year CSS(training cohort: 0.48, 0.45, 0.52; validation cohort: 0.42, 0.37, 0.37), IDI for the 3-, 6-, and 9-year CSS (training cohort: 0.09, 0.10, 0.11; validation cohort: 0.07, 0.08, 0.08). The Kaplan-Meier curve revealed that the new risk classification system possesses a more extraordinary ability to identify patients in different risk groups than the AJCC staging. Conclusion: A practical prognostic nomogram and novel risk classification system have been developed to efficiently predict the prognosis of EO-LARC. These tools can serve as a guide to individualize patient treatment and improve clinical decision-making.

A novel enemy of cancer: recent investigations into protozoan anti-tumor properties.

Cancer remains a significant global health issue, despite advances in screening and treatment. While existing tumor treatment protocols such as surgery, chemotherapy, radiotherapy, targeted therapy, and immunotherapy have proven effective in enhancing the prognosis for some patients, these treatments do not benefit all patients. Consequently, certain types of cancer continue to exhibit a relatively low 5-year survival rate. Therefore, the pursuit of novel tumor intervention strategies may help improve the current effectiveness of tumor treatment. Over the past few decades, numerous species of protozoa and their components have exhibited anti-tumor potential via immune and non-immune mechanisms. This discovery introduces a new research direction for the development of new and effective cancer treatments. Through in vitro experiments and studies involving tumor-bearing mice, the anti-tumor ability of Toxoplasma gondii, Plasmodium, Trypanosoma cruzi, and other protozoa have unveiled diverse mechanisms by which protozoa combat cancer, demonstrating encouraging prospects for their application. In this review, we summarize the anti-tumor ability and anti-tumor mechanisms of various protozoa and explore the potential for their clinical development and application.

Plasmodium Circumsporozoite Protein Enhances the Efficacy of Gefitinib in Lung Adenocarcinoma Cells by Inhibiting Autophagy via Proteasomal Degradation of LC3B.

Background: Almost all lung adenocarcinoma (LUAD) patients with EGFR mutant will develop resistance to EGFR-TKIs, which limit the long-term clinical application of these agents. Accumulating evidence shows one of the main reasons for resistance to EGFR-TKIs is induction of autophagy in tumor cells. Our previous study found that circumsporozoite protein (CSP) in Plasmodium can suppress autophagy in host hepatocytes. However, it is unknown whether CSP-mediated inhibition of autophagy could improve the anti-tumor effect of EGFR-TKIs. Methods: We constructed A549 and H1975 cell lines with stable overexpression of CSP (OE-CSP cells). CCK-8, Lactate Dehydrogenase (LDH), flow cytometry, and colony analysis were performed to observe the effect of CSP overexpression on cell viability, apoptosis rate, and colony formation ratio. The sensitizing effect of CSP on gefitinib was evaluated in vivo using a subcutaneous tumor model in nude mice and immunohistochemical assay. The role of CSP in regulation of autophagy was investigated by laser confocal microscopy assay and western blotting. A transcriptome sequencing assay and real-time polymerase chain reaction were used to determine the levels of mRNA for autophagy-related proteins. Cycloheximide (CHX), MG132, TAK-243, and immunoprecipitation assays were used to detect and confirm proteasomal degradation of LC3B. Results: OE-CSP A549 and H1975 cells were more sensitive to gefitinib, demonstrating significant amounts of apoptosis and decreased viability. In the OE-CSP group, autophagy was significantly inhibited, and there was a decrease in LC3B protein after exposure to gefitinib. Cell viability and colony formed ability were recovered when OE-CSP cells were exposed to rapamycin. In nude mice with xenografts of LUAD cells, inhibition of autophagy by CSP resulted in suppression of cell growth, and more marked apoptosis during exposure to gefitinib. CSP promoted ubiquitin-proteasome degradation of LC3B, leading to inhibition of autophagy in LUAD cells after treatment with gefitinib. When LUAD cells were treated with ubiquitin activating enzyme inhibitor TAK-243, cell viability, apoptosis, and growth were comparable between the OE-CSP group and a control group both in vivo and in vitro. Conclusion: CSP can inhibit gefitinib-induced autophagy via proteasomal degradation of LC3B, which suggests that CSP could be used as an autophagy inhibitor to sensitize EGFR-TKIs.

Repositioning of Antiparasitic Drugs for Tumor Treatment.

Drug repositioning is a strategy for identifying new antitumor drugs; this strategy allows existing and approved clinical drugs to be innovatively repurposed to treat tumors. Based on the similarities between parasitic diseases and cancer, recent studies aimed to investigate the efficacy of existing antiparasitic drugs in cancer. In this review, we selected two antihelminthic drugs (macrolides and benzimidazoles) and two antiprotozoal drugs (artemisinin and its derivatives, and quinolines) and summarized the research progresses made to date on the role of these drugs in cancer. Overall, these drugs regulate tumor growth via multiple targets, pathways, and modes of action. These antiparasitic drugs are good candidates for comprehensive, in-depth analyses of tumor occurrence and development. In-depth studies may improve the current tumor diagnoses and treatment regimens. However, for clinical application, current investigations are still insufficient, warranting more comprehensive analyses.

Moesin as a prognostic indicator of lung adenocarcinoma improves prognosis by enhancing immune lymphocyte infiltration.

BACKGROUND: Ezrin-radixin-moesin (ERM) have been explored in many cancer processes. Moesin, as its component, has also been found to play an important role in the prognosis of cancer patients, tumor metastasis, drug resistance, and others. Especially in regulating the immunity, but most results came from direct studies on immune cells, there is no clear conclusion on whether moesin has similar effects in tumor cells. And moesin has certain research results in many cancers in other aspects, but there are few about moesin in lung adenocarcinoma (LUAD). METHODS: We detect the expression of moesin in 82 LUAD and matched normal tissue samples by immunohistochemistry. Besides, for the pathological feature, we did a detailed statistical analysis. And with the help of various databases, we have done in-depth exploration of moesin's ability to enhance the extent of immune lymphocyte infiltration. RESULTS: Moesin is a poor expression in lung cancer tissues than the corresponding normal samples. And this phenomenon had a strongly associated with the prognosis and TNM stage of these LUAD patients. Moesin can enhance the infiltration of multiple immune lymphocytes in lung cancer. And this may be related to the interaction between moesin and various inflammatory molecules. CONCLUSIONS: Moesin is a newly index for the prognosis of LUAD and improves the prognosis of LUAD patients by regulating a variety of inflammation-related molecules to enhance immune lymphocytes infiltration.

Effect analysis of intradermal hyaluronic acid injection to treat enlarged facial pores.

OBJECTIVE: To investigate the clinical application and efficacy of intradermal injection of low molecular weight hyaluronic acid (LMW-HA) for treating enlarged facial pores. METHODS: From January 2015 to May 2016, 42 subjects who sought aesthetic treatment underwent intradermal injection of LMW-HA to improve enlarged facial pores. For each treatment, 2.5 mL (25 mg) of LMW-HA was injected into the skin of the full face. The treatment was repeated 2-5 times with an interval of 1 to 1.5 months between consecutive treatments. The postoperative follow-up period was 1 to 6 months. Statistical analysis was used to compare the degree of enlargement of facial pores before and after injection. The clinical efficacy and adverse effects were recorded. RESULTS: The enlarged facial pores before and after treatment were categorized and subjected to the Wilcoxon matched-pairs signed-rank test. The difference was statistically significant (P<.01). The improvement rate was 40.03±18.41%. No infection, nodules, or pigmentation was reported at the injection sites in the subjects who sought aesthetic treatment. The overall satisfaction rate was 92.8%. CONCLUSION: Intradermal injection of LMW-HA can significantly improve skin texture, reduce pore size, and enhance skin radiance. The injection technique was simple, safe, and effective and could easily be extended to clinical practice.

Mesenchymal stem cells promote colorectal cancer progression through AMPK/mTOR-mediated NF-κB activation.

Mesenchymal stem cells (MSCs) exert a tumor-promoting effect in a variety of human cancers. This study was designed to identify the molecular mechanisms related to the tumor-promoting effect of MSCs in colorectal cancer. In vitro analysis of colorectal cancer cell lines cultured in MSC conditioned media (MSC-CM) showed that MSC-CM significantly promoted the progression of the cancer cells by enhancing cell proliferation, migration and colony formation. The tumorigenic effect of MSC-CM was attributed to altered expression of cell cycle regulatory proteins and inhibition of apoptosis. Furthermore, MSC-CM induced high level expression of a number of pluripotency factors in the cancer cells. ELISAs revealed MSC-CM contained higher levels of IL-6 and IL-8, which are associated with the progression of cancer. Moreover, MSC-CM downregulated AMPK mRNA and protein phosphorylation, but upregulated mTOR mRNA and protein phosphorylation. The NF-κB pathway was activated after addition of MSC-CM. An in vivo model in Balb/C mice confirmed the ability of MSC-CM to promote the invasion and proliferation of colorectal cancer cells. This study indicates that MSCs promote the progression of colorectal cancer via AMPK/mTOR-mediated NF-κB activation.

Short-term memory of danger signals or environmental stimuli in mesenchymal stem cells: implications for therapeutic potential.

Mesenchymal stem/stromal cells (MSCs) possess some characteristics of immune cells, including a pro-inflammatory phenotype, an immunosuppressive phenotype, antibacterial properties and the expression of Toll-like receptor proteins. Here we show that, similar to immune cells, MSCs retain information from danger signals or environmental stimuli for a period of time. When treated with the pro-inflammatory factors lipopolysaccharide (LPS) or tumor necrosis factor-α (TNF-α), MSCs display increased expression of IL-6, IL-8 and MCP-1. Following re-plating and several rounds of cell division in the absence of stimulating factors, the expression of IL-6, IL-8 and MCP-1 remained higher than in untreated cells for over 7 days. A spike in cytokine secretion occurred when cells were exposed to a second round of stimulation. We primed MSCs with LPS and LPS-primed MSCs had better therapeutic efficacy at promoting skin flap survival in a diabetic rat model than did unprimed MSCs. Finally, we found that several microRNAs, including miR146a, miR150 and miR155, along with the modification of DNA by 5-hydroxymethylcytosine (5hmC), mediate the MSC response to LPS and TNF-α stimulation. Collectively, our data suggest that MSCs have a short-term memory of environmental signals, which may impact their therapeutic potential.

Effect of serum choice on replicative senescence in mesenchymal stromal cells.

BACKGROUND AIMS: Multipotent mesenchymal stromal cells (MSCs) are promising candidates for innovative cell therapeutic applications. Before their use, however, they usually need to be expanded in vitro with serum-supplemented media. MSCs can undergo replicative senescence during in vitro expansion, but it is not yet clear how serum supplements influence this process. METHODS: In the present study, we compared how media supplemented with fetal bovine serum (FBS) or calf serum (CS) affected morphology, proliferation, differentiation, senescence and other functional characteristics of human umbilical cord-derived MSCs (UC-MSCs). RESULTS: UC-MSCs cultured in both FBS- and CS-containing media were able to differentiate along osteogenic and adipogenic lineages but ultimately reached proliferation arrest. However, senescence-associated characteristics, such as ß-galactosidase activity, reactive oxygen species levels, proliferation rate and gene expression, demonstrate that UC-MSCs grown with FBS have better proliferation potential and differentiation capacity. In contrast, UC-MSCs grown with CS have a higher proportion of apoptotic cells and senescent characteristics. Possible mechanisms for the observed phenotypes include changes in gene expression (Bax, p16, p21 and p53) and cytokine production (interleukin-6 and interleukin-8). CONCLUSIONS: This study demonstrates that FBS-supplemented media provides a better microenvironment for the expansion of UC-MSCs in vitro than CS-supplemented media. This work provides insight into MSCs generation practices for use in basic research and clinical therapies.

Adipose-derived stem cells cooperate with fractional carbon dioxide laser in antagonizing photoaging: a potential role of Wnt and ß-catenin signaling.

BACKGROUND: It is well established that adipose-derived stem cells (ADSCs) produce and secrete cytokines/growth factors that antagonize UV-induced photoaging of skin. However, the exact molecular basis underlying the anti-photoaging effects exerted by ADSCs is not well understood, and whether ADSCs cooperate with fractional carbon dioxide (CO2) laser to facilitate photoaging skin healing process has not been explored. Here, we investigated the impacts of ADSCs on photoaging in a photoaging animal model, its associated mechanisms, and its functional cooperation with fractional CO2 laser in treatment of photoaging skin. RESULTS: We showed that ADSCs improved dermal thickness and activated the proliferation of dermal fibroblast. We further demonstrated that the combined treatment of ADSCs and fractional CO2 laser, the latter which is often used to resurface skin and treat wrinkles, had more beneficial effects on the photoaging skin compared with each individual treatment. In our prepared HDF photoaging model, flow cytometry showed that, after adipose derived stem cells conditioned medium (ADSC-CM) co-cultured HDF photoaging model, the cell proliferation rate is higher than UVB irradiation induced HDF modeling (p < 0.05). Additionally, the expressions of ß-catenin and Wnt3a, which were up-regulated after the transplantation of ADSCs alone or in combination with fractional CO2 laser treatment. And the expression of wnt3a and ß-catenin has the positive correlation with photoaging related protein TGF-ß2 and COLI. We also verified these protein expressions in tissue level. In addition, after injected SFRP2 into ADSC-CM co-cultured HDF photoaging model, wnt3a inhibitor, compared with un-intervened group, wnt3a, ß-catenin protein level significantly decreased. CONCLUSION: Both ADSCs and fractional CO2 laser improved photoaging skin at least partially via targeting dermal fibroblast activity which was increased in photoaging skin. The combinatorial use of ADSCs and fractional CO2 laser synergistically improved the healing process of photoaging skin. Thus, we provide a strong rationale for a combined use of ADSCs and fractional CO2 laser in treatment of photoaging skin in clinic in the future. Moreover, we provided evidence that the Wnt/ß-catenin signaling pathway may contribute to the activation of dermal fibroblast by the transplantation of ADSCs in both vitro and vivo experiment.

Human chorionic gonadotropin decreases human breast cancer cell proliferation and promotes differentiation.

Human chorionic gonadotropin (hCG) is a glycoprotein produced by placental trophoblasts. Previous studies indicated that hCG could be responsible for the pregnancy-induced protection against breast cancer in women. It is reported that hCG decreases proliferation and invasion of breast cancer MCF-7 cells. Our research also demonstrates that hCG can reduce the proliferation of MCF-7 cells by downregulating the expression of proliferation markers, proliferating cell nuclear antigen (PCNA), and proliferation-related Ki-67 antigen (Ki-67). Interestingly, we find here that hCG elevates the state of cellular differentiation, as characterized by the upregulation of differentiation markers, ß-casein, cytokeratin-18 (CK-18), and E-cadherin. Inhibition of hCG secretion or luteinizing hormone/hCG receptors (LH/hCGRs) synthesis can weaken the effect of hCG on the induction of cell differentiation. Furthermore, hCG can suppress the expression of estrogen receptor alpha. hCG activated receptor-mediated cyclic adenosine monophosphate/protein kinase A signaling pathway. These findings indicated that a protective effect of hCG against breast cancer may be associated with its growth inhibitory and differentiation induction function in breast cancer cells.

Re-expression and epigenetic modification of maspin induced apoptosis in MCF-7 cells mediated by myocardin.

Breast cancer is the leading cause of cancer death in women worldwide. It is well known that oncogene activation and anti-oncogene inactivation affect the development and progression of breast cancer, but the role of oncogene activation and anti-oncogene inactivation in breast cancer is still not fully understood. We now report that maspin acts as a tumor suppressor gene to induce MCF-7 cell apoptosis. In addition, maspin promoter hypermethylation and histone hypoacetylation lead to silencing of maspin gene expression in MCF-7 cells. Moreover, DNA methyltransferase (DNMT) inhibitor 5-aza-2'-deoxycytidine (5-aza-dc) and/or the histone deacetylase (HDAC) inhibitor Trichostatin A (TSA) strongly up-regulated the expression of maspin in MCF-7 cells. Notably, myocardin can promote the re-expression of maspin in MCF-7 cells. Luciferase assay shows that myocardin activates the transcription of maspin promoter by CArG box. More importantly, 5-aza-dc/TSA and myocardin synergetically enhance re-expression of maspin and augment maspin-mediated apoptosis in MCF-7 cells. Thus, these data reveal the new insight that myocardin meditates apoptosis in breast cancer through affecting maspin re-expression and epigenetic modification to regulate the development of breast cancer, thereby raising the possibility of its use in breast cancer therapy.

Estrogen receptor α mediates proliferation of breast cancer MCF-7 cells via a p21/PCNA/E2F1-dependent pathway.

High expression of estrogen receptor α (ERα) is associated with a poor prognosis that correlates closely with cellular proliferation in breast cancer. However, the exact molecular mechanism by which ERα controls breast cancer cell proliferation is not clear. Here we report that ERα regulates the cell cycle by suppressing p53/p21 and up-regulating proliferating cell nuclear antigen (PCNA) and proliferation-related Ki-67 antigen (Ki-67) to promote proliferation of MCF-7 cells. In addition, 17-ß-estradiol (E2) enhances ERα-induced proliferation of MCF-7 cells by stimulating expression of PCNA and Ki-67. Knockdown of ERα significantly affects PCNA/Ki-67 and p53/p21 expression. Furthermore, ERα inhibits the transcriptional activity of p53/p21 in an estrogen response element-dependent manner. More importantly, we provide new evidence that ERα mediates proliferation of MCF-7 cells by up-regulating miR-17 to silence the expression of p21. Thus, these data provide new insights into the underlying effect of ERα on breast cancer proliferation.