Current Applications of Colorectal Cancer Organoids: A Review.

Colorectal cancer is a prevalent malignancy, with advanced and metastatic forms exhibiting poor treatment outcomes and high relapse rates. To enhance patient outcomes, a comprehensive understanding of the pathophysiological processes and the development of targeted therapies are imperative. The high heterogeneity of colorectal cancer demands precise and personalized treatment strategies. Colorectal cancer organoids, a three-dimensional in vitro model, have emerged as a valuable tool for replicating tumor biology and exhibit promise in scientific research, disease modeling, drug screening, and personalized medicine. In this review, we present an overview of colorectal cancer organoids and explore their applications in research and personalized medicine, while also discussing potential future developments in this field.

Association of Overweight and Inflammatory Indicators with Breast Cancer: A Cross-Sectional Study in Chinese Women.

Objective: This cross-sectional study aimed to explore the association of overweight and inflammatory indicators with breast cancer risk in Chinese patients. Methods: Weight, height, and peripheral blood inflammatory indicators, including white blood cell count (WBC), neutrophil count (NE), lymphocyte count (LY), platelet count (PLT) and the concentration of hypersensitivity C-reactive protein (hsCRP), were collected in 383 patients with benign breast lumps (non-cancer) and 358 patients with malignant breast tumors (cancer) at the First Affiliated Hospital of Soochow University, China, from March 2018 to July 2020. Body mass index (BMI), neutrophil-lymphocyte ratio (NLR), platelet-lymphocyte ratio (PLR) and systemic immune-inflammation index (SII) were determined according to the ratio equation. The correlations among overweight, inflammatory indicators, and the proportion of non-cancer or cancer cases were analyzed. Results: BMI is associated with an increased breast cancer risk. Compared with non-cancer patients, the average WBC count, NE count, NLR, and level of hsCRP were significantly higher in cancer patients. The level of hsCRP was closely associated with the size of malignant breast tumors. Conclusion: We conclude that overweight and high levels of hsCRP may serve as putative risk factors for malignant breast tumors in Chinese women.

[Effect of Huangqin Decoction on pyroptosis pathway of NLRP3/caspase-1 in mice with ulcerative colitis].

To explore the effect of Huangqin Decoction on ulcerative colitis(UC) pyroptosis, and to explain the mechanism of pyroptosis based on NOD-like receptor thermoprotein domain 3(NLRP3)/cysteine proteinase 1(caspase-1) pathway. The animal model of UC induced with 3% dextran sodium sulfate(DSS) was established. The experimental animals were divided into control group, model group, low-dose(4.55 g·kg~(-1)), medium-dose(9.1 g·kg~(-1)) and high-dose(18.2 g·kg~(-1)) Huangqin Decoction groups and salazosulfapyridine group(0.45 g·kg~(-1)). While modeling, intragastric administration was given for 7 consecutive days. On the 8 th day, the mice were euthanized, the colon length was collected, and the histopathological changes were observed by HE staining. The content of interleukin-18(IL-18) was observed by ELISA. The content of lactatedehydrogenase(LDH)was determined by microplate method. TUNEL assay kit was used to detect the cell death. The immunohistochemical staining was used to detect the expressions of NLRP3 and apoptosis-associated speck-like protein containing a CARD(ASC). Western blot was used to detect the expressions of interleukin-1ß(IL-1ß), caspase-1 and gasdermin D(GSDMD).The experimental study showed that compared with normal group, the LDH content, TUNEL positive staining, inflammatory factors(IL-18, IL-1ß), and proteins associated with pyroptosis were significantly increased(P<0.05). Compared with model control group, the LDH content, TUNEL positive staining, inflammatory factors(IL-18, IL-1ß), and proteins associated with pyroptosis were decreased, and these results were more significant in high-dose groups(P<0.05). The results of HE staining showed that Huangqin Decoction could improve the pathological changes of colon. Huangqin Decoction could inhibit UC cell pyroptosis, and the mechanism may be closely related to NLRP3/caspase-1 signaling pathway.

Intracellular MLCK1 diversion reverses barrier loss to restore mucosal homeostasis.

Epithelial barrier loss is a driver of intestinal and systemic diseases. Myosin light chain kinase (MLCK) is a key effector of barrier dysfunction and a potential therapeutic target, but enzymatic inhibition has unacceptable toxicity. Here, we show that a unique domain within the MLCK splice variant MLCK1 directs perijunctional actomyosin ring (PAMR) recruitment. Using the domain structure and multiple screens, we identify a domain-binding small molecule (divertin) that blocks MLCK1 recruitment without inhibiting enzymatic function. Divertin blocks acute, tumor necrosis factor (TNF)-induced MLCK1 recruitment as well as downstream myosin light chain (MLC) phosphorylation, barrier loss, and diarrhea in vitro and in vivo. Divertin corrects barrier dysfunction and prevents disease development and progression in experimental inflammatory bowel disease. Beyond applications of divertin in gastrointestinal disease, this general approach to enzymatic inhibition by preventing access to specific subcellular sites provides a new paradigm for safely and precisely targeting individual properties of enzymes with multiple functions.

In Vitro and In Vivo Approaches to Determine Intestinal Epithelial Cell Permeability.

The intestinal barrier defends against pathogenic microorganism and microbial toxin. Its function is regulated by tight junction permeability and epithelial cell integrity, and disruption of the intestinal barrier function contributes to progression of gastrointestinal and systemic disease. Two simple methods are described here to measure the permeability of intestinal epithelium. In vitro, Caco-2BBe cells are plated in tissue culture wells as a monolayer and transepithelial electrical resistance (TER) can be measured by an epithelial (volt/ohm) meter. This method is convincing because of its user-friendly operation and repeatability. In vivo, mice are gavaged with 4 kDa fluorescein isothiocyanate (FITC)-dextran, and the FITC-dextran concentrations are measured in collected serum samples from mice to determine the epithelial permeability. Oral gavage provides an accurate dose, and therefore is the preferred method to measure the intestinal permeability in vivo. Taken together, these two methods can measure the permeability of the intestinal epithelium in vitro and in vivo, and hence be used to study the connection between diseases and barrier function.

Myosin regulatory light chain phosphorylation is associated with leiomyosarcoma development.

Leiomyosarcoma is a rare malignant smooth muscle tumor which can be very unpredictable. Myosin II is involved in many functions, including cell contraction, migration, and adhesion. The phosphorylation of myosin regulatory light chain (MLC) by myosin light chain kinase (MLCK) determines the activity of Myosin II. However, it is still unclear whether MLC phosphorylation is involved in cell proliferation in leiomyosarcoma. In this study, we aimed to explore the role of MLCK-dependent MLC phosphorylation in leiomyosarcoma development. We found that the expression of MLCK, phosphorylated MLC, and Ki67 in leiomyosarcoma was significantly higher than in leiomyoma and adjacent normal smooth muscle cells. MLCK expression was significantly correlated with phosphorylated MLC level. Kaplan-Meier survival analysis revealed that patients with high expression of MLCK or phosphorylated MLC had shorter overall survival times compared with the patients with low expression of MLCK or phosphorylated MLC. In vitro studies revealed a causative link between MLC phosphorylation and cellular proliferation as expression of phosphomimetic MLC (T19D, S20D) increased cellular proliferation as assessed by Ki67 staining. In contrast, MLCK specific inhibitor reduced cellular proliferation. We concluded that MLCK, phosphorylated MLC and Ki67 were overexpressed in leiomyosarcoma. MLCK dependent MLC phosphorylation might be responsible for the high proliferative state in leiomyosarcoma. MLCK and phosphorylated MLC are potential prognostic indicators of leiomyosarcoma.

Characterization of isoform expression and subcellular distribution of MYPT1 in intestinal epithelial cells.

The regulation of intestinal epithelial permeability requires phosphorylation of myosin regulatory light chain (MLC). The phosphorylation status of MLC is regulated by myosin light chain phosphatase (MLCP) activities. The activity of the catalytic subunit of MLCP (PP1cδ) toward MLC depends on its regulatory subunit (MYPT1). In this study, we revealed the presence of two MYPT1 isoforms, full length and variant 2 in human intestinal (Caco-2) epithelial cells and isolated intestinal epithelial cells (IECs) from mice. In confluent Caco-2 cells, MYPT1 was distributed at cell-cell contacts and colocalized with F-actin. These results suggest that MYPT1 isoforms are expressed in intestinal epithelial cells and MYPT1 may be involved in the regulation of intestinal epithelial barrier function.

The molecular mechanism of CRISPR/Cas9 system and its application in gene therapy of human diseases.

CRISPR/Cas system is an adaptive immune system that confers resistance to exogenous virus or plasmid in bacteria and archaea. In recent years, the booming CRISPR/Cas9 genome editing technology modified from type2 CRISPR/Cas adaptive immune system has been widely applied to various research fields of life science and led to revolutionary changes. In this review, we summarize the origin and development of CRISPR/Cas9 genome editing technology as well as its applications in life science research. We focus on the latest application of this system in gene therapy of human diseases and the associated side/off-target effects, which may provide references for researchers in related areas.