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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.
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Neoplasias Colorrectales , Organoides , Medicina de Precisión , Humanos , Organoides/patología , Neoplasias Colorrectales/patología , Neoplasias Colorrectales/terapia , AnimalesRESUMEN
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.
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Falling film evaporation technology is widely used in the treatment of salt-containing wastewater in coal chemical industry. However, there is still a lack of research on the inlet method of vertical falling film evaporation tubes. In this paper, the heat and mass transfer processes of saline wastewater under vertical and tangential inlets were investigated using numerical simulations. On this basis, the differences in flow and heat transfer processes between saline wastewater and pure water under tangential inlet were investigated. The results showed that the flow velocity of saline wastewater with a falling film evaporation tube in a tangential inlet mode was larger. Meanwhile, the turbulence in this way was more intense and the fluid temperature in the vertical tube was higher. Saline wastewater has higher temperature and smaller liquid volume fraction than pure water liquid membrane in the range of 193-1,000 mm from the inlet. The use of tangential inlet method to treat salt-containing wastewater has higher evaporation efficiency and is a very effective way to guide the improvement of heat transfer efficiency.
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Calor , Aguas Residuales , Bahías , TemperaturaRESUMEN
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.
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Colitis Ulcerosa , Piroptosis , Animales , Caspasa 1/genética , Colitis Ulcerosa/tratamiento farmacológico , Ratones , Proteína con Dominio Pirina 3 de la Familia NLR/genética , Scutellaria baicalensisRESUMEN
Data GSE75214 and GSE48959 that contained ulcerative colitis(UC) in the active stage was download from GEO database. Differential genes of UC in the active phase were obtained by using adjusted P<0.05 and |log_2 FC|≥1.5, which was the screening criteria. PPI analysis was performed in the STRING database, and GO and KEGG pathway analysis was performed in DAVID database. Cytoscape was used to visualize differential genes, and calculate key genes in the active phase. Coremine Medical was used to analyze and systematically evaluate traditional Chinese medicines for treating key genes. Finally, 139 differentially expressed genes in the active phase were screened out, which included the 109 up-regulated genes and 30 down-regulated genes. DAVID analyzed that the biology and pathways of these differential genes were mainly concentrated in inflammatory response, immune response, chemokine activity, TNF pathway, NF-κB pathway, and Toll-like receptor pathway. Cytoscape software calculated that IL-6, CXCL8, IL-1ß, MMP9, CXCL1, ICAM1, CXCL10, TIMP1, PTGS2 and CXCL9 were the key genes of UC in the active phase. According to Coremine Medical analysis, traditional Chinese medicines for UC in the active stage included Curcumae Longae Rhizoma, Scutellariae Radix, Curcumae Radix had clearing heat clearing damp, reducing fire and detoxifying effects, which was in line with the pathogenesis of UC active stage, and was often used in clinical treatment of dampness-heat diarrhea. Therefore, Huangqin Decoction, which Scutellariae Radix was the principal drug, was selected for systematic evaluation. The evaluation showed that Scutellariae Radix was superior to Western medicine in terms of improving clinical efficiency, reducing inflammatory factors and immunoglobulin levels, with statistically significant differences and fewer adverse reactions. This study provided a new idea for further research on the pathogenesis of UC in the active phase by analyzing the genes and their mechanism of action, and the systematic evaluation of Chinese medicine for the treatment of UC active stage provided a basis for the clinical prevention and treatment of UC by Chinese medicine.
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Colitis Ulcerosa , Medicamentos Herbarios Chinos , Biología Computacional , Humanos , Medicina Tradicional China , Scutellaria baicalensisRESUMEN
The intestinal epithelium acts as a barrier that prevents luminal contents, such as pathogenic microbiota and toxins, from entering the rest of the body. Epithelial barrier function requires the integrity of intestinal epithelial cells. While epithelial cell proliferation maintains a continuous layer of cells that forms a barrier, epithelial damage leads to barrier dysfunction. As a result, luminal contents can across the intestinal barrier via an unrestricted pathway. Dysfunction of intestinal barrier has been associated with many intestinal diseases, such as inflammatory bowel disease. Isolated mouse intestinal crypts can be cultured and maintained as crypt-villus-like structures, which are termed intestinal organoids or "enteroids". Enteroids are ideal to study the proliferation and cell death of intestinal epithelial cells in vitro. In this protocol, we describe a simple method to quantify the number of proliferative and dead cells in cultured enteroids. 5-ethynyl-2'-deoxyuridine (EdU) and propidium iodide are used to label proliferating and dead cells in enteroids, and the proportion of proliferating and dead cells are then analyzed by flow cytometry. This is a useful tool to test the effects of drug treatment on intestinal epithelial cell proliferation and cell survival.
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Mucosa Intestinal/metabolismo , Animales , Proliferación Celular , Citometría de Flujo , RatonesRESUMEN
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.
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Homeostasis , Mucosa Intestinal/metabolismo , Espacio Intracelular/enzimología , Quinasa de Cadena Ligera de Miosina/metabolismo , Actomiosina/metabolismo , Animales , Células CACO-2 , Enfermedad Crónica , Homeostasis/efectos de los fármacos , Humanos , Inflamación/patología , Enfermedades Inflamatorias del Intestino/patología , Mucosa Intestinal/efectos de los fármacos , Yeyuno/efectos de los fármacos , Yeyuno/metabolismo , Yeyuno/patología , Ratones , Cadenas Ligeras de Miosina/metabolismo , Quinasa de Cadena Ligera de Miosina/química , Fosforilación/efectos de los fármacos , Dominios Proteicos , Bibliotecas de Moléculas Pequeñas/farmacología , Uniones Estrechas/efectos de los fármacos , Uniones Estrechas/metabolismo , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
BACKGROUND & AIMS: Epithelial regeneration is essential for homeostasis and repair of the mucosal barrier. In the context of infectious and immune-mediated intestinal disease, interleukin (IL) 22 is thought to augment these processes. We sought to define the mechanisms by which IL22 promotes mucosal healing. METHODS: Intestinal stem cell cultures and mice were treated with recombinant IL22. Cell proliferation, death, and differentiation were assessed in vitro and in vivo by morphometric analysis, quantitative reverse transcriptase polymerase chain reaction, and immunohistochemistry. RESULTS: IL22 increased the size and number of proliferating cells within enteroids but decreased the total number of enteroids. Enteroid size increases required IL22-dependent up-regulation of the tight junction cation and water channel claudin-2, indicating that enteroid enlargement reflected paracellular flux-induced swelling. However, claudin-2 did not contribute to IL22-dependent enteroid loss, depletion of Lgr5+ stem cells, or increased epithelial proliferation. IL22 induced stem cell apoptosis but, conversely, enhanced proliferation within and expanded numbers of transit-amplifying cells. These changes were associated with reduced wnt and notch signaling, both in vitro and in vivo, as well as skewing of epithelial differentiation, with increases in Paneth cells and reduced numbers of enteroendocrine cells. CONCLUSIONS: IL22 promotes transit-amplifying cell proliferation but reduces Lgr5+ stem cell survival by inhibiting notch and wnt signaling. IL22 can therefore promote or inhibit mucosal repair, depending on whether effects on transit-amplifying or stem cells predominate. These data may explain why mucosal healing is difficult to achieve in some inflammatory bowel disease patients despite markedly elevated IL22 production.
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Interleucinas/farmacología , Receptores Acoplados a Proteínas G/metabolismo , Receptores Notch/metabolismo , Células Madre/citología , Células Madre/metabolismo , Vía de Señalización Wnt/efectos de los fármacos , Animales , Apoptosis/efectos de los fármacos , Biomarcadores/metabolismo , Recuento de Células , Diferenciación Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Claudina-2/metabolismo , Enterocitos/citología , Enterocitos/efectos de los fármacos , Enterocitos/metabolismo , Intestinos/citología , Ratones , Ratones Endogámicos C57BL , Organoides/metabolismo , Células Madre/efectos de los fármacos , Regulación hacia Arriba/efectos de los fármacos , Interleucina-22RESUMEN
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.
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Células Epiteliales/química , Mucosa Intestinal/química , Animales , Humanos , Mucosa Intestinal/patología , Ratones , PermeabilidadRESUMEN
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.
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Carcinogénesis/metabolismo , Carcinogénesis/patología , Leiomiosarcoma/metabolismo , Leiomiosarcoma/patología , Cadenas Ligeras de Miosina/metabolismo , Línea Celular Tumoral , Proliferación Celular , Femenino , Humanos , Estimación de Kaplan-Meier , Antígeno Ki-67/metabolismo , Persona de Mediana Edad , Miocitos del Músculo Liso/metabolismo , Miocitos del Músculo Liso/patología , Quinasa de Cadena Ligera de Miosina/metabolismo , Fosforilación , PronósticoRESUMEN
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.
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Empalme Alternativo , Fosfatasa de Miosina de Cadena Ligera/genética , Actinas/metabolismo , Animales , Células CACO-2 , Línea Celular Tumoral , Células Cultivadas , Células Epiteliales/metabolismo , Humanos , Ratones , Ratones Endogámicos C57BL , Especificidad de Órganos , Isoformas de Proteínas/genéticaRESUMEN
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.
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Inmunidad Adaptativa/genética , Sistemas CRISPR-Cas , Ingeniería Genética/métodos , Terapia Genética/métodos , Endonucleasas/genética , Endonucleasas/metabolismo , Ingeniería Genética/tendencias , Terapia Genética/tendencias , Infecciones por VIH/genética , Infecciones por VIH/inmunología , Infecciones por VIH/terapia , Humanos , Modelos Genéticos , Neoplasias/genética , Neoplasias/inmunología , Neoplasias/terapia , ARN Guía de Kinetoplastida/genética , ARN Guía de Kinetoplastida/metabolismoRESUMEN
<p><b>OBJECTIVE</b>To observe the effect of mini-implant lengths on stress distribution in peri-implant surface.</p><p><b>METHODS</b>The 3D finite element analysis mandible and mini-implant models with diameter of 1.6 mm, lengths of 6, 8, 10 and 12 mm were established. The mini-implants were inserted into designed site of mandibular vertically, respectively. A force of 1.96 N were applied mesioly and 45 degrees tilted mesio-vertically in models. The stress distribution under every condition was recorded and analyzed.</p><p><b>RESULTS</b>When load was applied mesially, the maximum stress varied from 3.500 to 3.765 MPa, the maximum displacement varied from 1.266 to 1.288 microm. When load was applied 45 degrees tilted mesio-vertically, the maximum stress varied from 4.075 to 4.510 MPa, the maximum displacement varied from 1.668 to 1.694 microm. All of the maximum stress and displacement of loading mesially were lower than loading mesio-vertically.</p><p><b>CONCLUSION</b>The change of the mini-implant length within 6-12 mm don't show much influence on the stress distribution. The loading type is an important factor influencing stress and displacement of peri-implant bone.</p>