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Fibroblast-Epithelium Co-culture Methods Using Epithelial Organoids and Cell Line-Derived Spheroids.Methods in Molecular Biology (Clifton,... 2024Fibroblasts are an integral cell type of mammary gland stroma, which plays crucial roles in development, homeostasis, and tumorigenesis of mammary epithelium....
Fibroblasts are an integral cell type of mammary gland stroma, which plays crucial roles in development, homeostasis, and tumorigenesis of mammary epithelium. Fibroblasts produce and remodel extracellular matrix proteins and secrete a plethora of paracrine signals, which instruct both epithelial and other stromal cells of the mammary gland through mechanisms, which have not been fully understood. To enable deciphering of the intricate fibroblast-epithelial interactions, we developed several 3D co-culture methods. In this chapter, we describe methods for establishment of various types of embedded 3D co-cultures of mammary fibroblasts with mammary epithelial organoids, mammary tumor organoids, or breast cancer spheroids to investigate the role of fibroblasts in mammary epithelial development, morphogenesis, and tumorigenesis. The co-culture types include dispersed, aggregated, and transwell cultures.
Topics: Animals; Humans; Coculture Techniques; Epithelial Cells; Mammary Glands, Animal; Epithelium; Cell Line; Fibroblasts; Organoids; Carcinogenesis
PubMed: 38393591
DOI: 10.1007/978-1-0716-3674-9_8 -
Anti-cancer Agents in Medicinal... 2024The aim of this study was to synthesize a library of novel di-sulfa drugs containing 1,3- diaryltriazene derivatives TS (1-13) by conjugation of diazonium salts of...
AIM
The aim of this study was to synthesize a library of novel di-sulfa drugs containing 1,3- diaryltriazene derivatives TS (1-13) by conjugation of diazonium salts of primary sulfonamides with sulfa drugs to investigate the cytotoxic effect of these new compounds in different cancer types and to determine their inhibitory activity against tumor-associated carbonic anhydrases IX and XII.
MATERIALS AND METHODS
A carbonic anhydrase inhibitory activity of the obtained compounds was evaluated against four selected human carbonic anhydrase isoforms (hCA I, hCA II, hCA IX and hCA XII) by a stoppedflow CO hydrase assay. In addition, , cytotoxicity studies were applied by using A549 (lung cancer), BEAS-2B (normal lung), MCF-7 (breast cancer), MDA-MB-231 (breast cancer), CRL-4010 (normal breast epithelium), HT-29 (colon cancer), and HCT -116 (colon cancer) cell lines.
RESULTS
As a result of the inhibition data, the 4-aminobenzenesulfonamide derivatives were more active than their 3-aminobenzenesulfonamide counterparts. More specifically, compounds TS-1 and TS-2, both of which have primary sulfonamides on both sides of the triazene linker, showed the best inhibitory activity against hCA IX with K values of 19.5 and 13.7 nM and also against hCA XII with K values of 6.6 and 8.3 nM, respectively. In addition, cytotoxic activity on the human breast cancer cell line MCF-7 showed that some derivatives of di-sulfa triazenes, such as TS-5 and TS-13, were more active than SLC-0111.
CONCLUSION
With the aim of developing more potent and isoform-selective CA inhibitors, these novel hybrid molecules containing sulfa drugs, triazene linkers, and the classical primary sulfonamide chemotype may be considered an interesting example of effective enzyme inhibitors and important anticancer agents.
Topics: Humans; Carbonic Anhydrase Inhibitors; Carbonic Anhydrase IX; Antineoplastic Agents; Carbonic Anhydrases; Cell Proliferation; Drug Screening Assays, Antitumor; Sulfonamides; Structure-Activity Relationship; Molecular Structure; Triazenes; Dose-Response Relationship, Drug; Antigens, Neoplasm
PubMed: 38362678
DOI: 10.2174/0118715206285326240207045249 -
DEN Open Apr 2024A 57-year-old woman with no significant medical history was referred after a colonoscopy for abdominal distension, which revealed a tumor in the lower rectum....
A 57-year-old woman with no significant medical history was referred after a colonoscopy for abdominal distension, which revealed a tumor in the lower rectum. Pre-operative colonoscopy showed the tumor was 12 mm in size, located from the anorectal junction to beyond the dentate line, and was diagnosed as high-grade intramucosal neoplasia or shallow submucosal invasive cancer. Endoscopic submucosal dissection was performed, and the lesion was resected en bloc. Pathological examination revealed moderately differentiated tubular adenocarcinoma with tubulovillous adenoma. The stratified squamous epithelium adjacent to the anal side of the lesion showed pagetoid spread of atypical cells with positive horizontal margins. We referred her to a surgeon for radical treatment. The mucosa surrounding the endoscopic submucosal dissection scar was normal on narrow-band imaging magnification. We marked its oral side endoscopically as the resected boundary. Transanal local excision was performed. The horizontal margins were positive because atypical cells had spread into the stratified squamous epithelium of the anorectal side of the lesion. The patient was followed on an outpatient basis. Sixty days postoperatively, residual tumor growth was observed. The second local resection was performed after mapping biopsy. All resection margins were negative, there was no lymphovascular invasion. One year after surgery, no recurrence was observed. Regarding endoscopic findings, there are no reports of endoscopic findings of the rectal mucosa, or the squamous epithelium of the anus of pagetoid spread. Here, we report a review of perianal Paget's Disease that resulted in difficulties in borderline diagnosis of pagetoid spread, resulting in multiple therapeutic interventions.
PubMed: 38343421
DOI: 10.1002/deo2.340 -
Cureus Jan 2024Background and objective Gynecomastia is a benign proliferation of ductal epithelium in the retroareolar region in male patients. The aim of this study was to...
Background and objective Gynecomastia is a benign proliferation of ductal epithelium in the retroareolar region in male patients. The aim of this study was to investigate the frequency of gynecomastia in male patients who underwent thoracic computed tomography (CT) imaging at our clinic, assess possible causes, highlight the imaging characteristics of gynecomastia, and compare our findings with the literature. Materials and methods Male patients over 18 years of age who underwent thoracic CT imaging in our clinic were included in the study. Patients were initially assessed based on age and the presence of gynecomastia. The patients with gynecomastia were evaluated in terms of age, gynecomastia localization (right, left, and bilateral), gynecomastia type (nodular, dentritic, and diffuse), and possible etiology. Results The study included 1500 patients with a mean age of 45.6±21.7 years, and 470 (31.3%) patients had gynecomastia. Gynecomastia was on the right side in 11.3%, on the left side in 11.1%, and bilateral in 77.7% of the patients. Gynecomastia was nodular in 52.1%, dendritic in 35.3%, and diffuse in 17.2% of the patients. The causative factor could not be identified in 44.3% of the patients with gynecomastia. Among cases where the etiology was identified (56.7%), the most common factors were cancer (23.4%), chronic kidney disease (CKD) (13.2%), and chronic hepatitis B (10.7%). Conclusion When evaluating thoracic CT, the breast area, in addition to the lungs, chest wall, and bone structures, should also be evaluated carefully. With the increased use of thoracic CT scans, incidentally detected gynecomastia in patients is also on the rise. Knowing the presence of gynecomastia is very important for the clinician to determine the etiology and treat the underlying disease. Therefore, detecting and reporting gynecomastia on thoracic CT can prevent unnecessary advanced breast imaging methods and play a very important role in treating the underlying etiology.
PubMed: 38304650
DOI: 10.7759/cureus.51509 -
Frontiers in Oncology 2023Several breast cancer (BC) patients showed urinary tract infection after adjuvant trastuzumab plus paclitaxel, but no case of urothelial injury has been reported. In...
Several breast cancer (BC) patients showed urinary tract infection after adjuvant trastuzumab plus paclitaxel, but no case of urothelial injury has been reported. In this case, we report a 47-year-old female patient with stage I invasive ductal carcinoma in the left breast presenting with urothelial injury after the combination of trastuzumab and paclitaxel. Initially, the patient was highly suspected of having urinary tract infection as she showed abdominal and low back pain, as well as urinary irritation symptoms and hematuria. Unfortunately, the conditions were not attenuated after anti-infection therapy. Contrast-enhanced CT showed extensive exudation and edema in the bilateral renal pelvis, ureter, and bladder, together with dilatation and effusion in the renal pelvis and ureter. Cystoscopy showed extensive congestion, edema, and erosion in the bladder epithelium. Pathological analysis demonstrated slight thinning or even loss in the uroepithelial cell layer and interstitial congestion. In addition, there was growth arrest in the epithelial cells. Immunohistochemistry indicated HER2 expression in the urothelial cells. Finally, the patient was diagnosed with urothelial injury after combination of paclitaxel and trastuzumab. The symptoms were spontaneously cured with no administration of any antibiotics in the 3-month follow-up.
PubMed: 38304030
DOI: 10.3389/fonc.2023.1258474 -
Cell Discovery Jan 2024Malignant forms of breast cancer refractory to existing therapies remain a major unmet health issue, primarily due to metastatic spread. A better understanding of the...
Malignant forms of breast cancer refractory to existing therapies remain a major unmet health issue, primarily due to metastatic spread. A better understanding of the mechanisms at play will provide better insights for alternative treatments to prevent breast cancer cell dispersion. Here, we identify the lysine methyltransferase SMYD2 as a clinically actionable master regulator of breast cancer metastasis. While SMYD2 is overexpressed in aggressive breast cancers, we notice that it is not required for primary tumor growth. However, mammary-epithelium specific SMYD2 ablation increases mouse overall survival by blocking the primary tumor cell ability to metastasize. Mechanistically, we identify BCAR3 as a genuine physiological substrate of SMYD2 in breast cancer cells. BCAR3 monomethylated at lysine K334 (K334me1) is recognized by a novel methyl-binding domain present in FMNLs proteins. These actin cytoskeleton regulators are recruited at the cell edges by the SMYD2 methylation signaling and modulate lamellipodia properties. Breast cancer cells with impaired BCAR3 methylation lose migration and invasiveness capacity in vitro and are ineffective in promoting metastases in vivo. Remarkably, SMYD2 pharmacologic inhibition efficiently impairs the metastatic spread of breast cancer cells, PDX and aggressive mammary tumors from genetically engineered mice. This study provides a rationale for innovative therapeutic prevention of malignant breast cancer metastatic progression by targeting the SMYD2-BCAR3-FMNL axis.
PubMed: 38296970
DOI: 10.1038/s41421-023-00644-x -
Clinical Proteomics Jan 2024Omics characterization of pancreatic adenocarcinoma tissue is complicated by the highly heterogeneous and mixed populations of cells. We evaluate the feasibility and...
BACKGROUND
Omics characterization of pancreatic adenocarcinoma tissue is complicated by the highly heterogeneous and mixed populations of cells. We evaluate the feasibility and potential benefit of using a coring method to enrich specific regions from bulk tissue and then perform proteogenomic analyses.
METHODS
We used the Biopsy Trifecta Extraction (BioTExt) technique to isolate cores of epithelial-enriched and stroma-enriched tissue from pancreatic tumor and adjacent tissue blocks. Histology was assessed at multiple depths throughout each core. DNA sequencing, RNA sequencing, and proteomics were performed on the cored and bulk tissue samples. Supervised and unsupervised analyses were performed based on integrated molecular and histology data.
RESULTS
Tissue cores had mixed cell composition at varying depths throughout. Average cell type percentages assessed by histology throughout the core were better associated with KRAS variant allele frequencies than standard histology assessment of the cut surface. Clustering based on serial histology data separated the cores into three groups with enrichment of neoplastic epithelium, stroma, and acinar cells, respectively. Using this classification, tumor overexpressed proteins identified in bulk tissue analysis were assigned into epithelial- or stroma-specific categories, which revealed novel epithelial-specific tumor overexpressed proteins.
CONCLUSIONS
Our study demonstrates the feasibility of multi-omics data generation from tissue cores, the necessity of interval H&E stains in serial histology sections, and the utility of coring to improve analysis over bulk tissue data.
PubMed: 38291365
DOI: 10.1186/s12014-024-09450-3 -
Microbiology Spectrum Mar 2024Despite the established concept of the human mammary gland (MG) as a habitat with its own microbiota, the exact mechanism of MG colonization is still elusive and a...
Despite the established concept of the human mammary gland (MG) as a habitat with its own microbiota, the exact mechanism of MG colonization is still elusive and a well-characterized model would reinforce studies of the MG microbiota development. We aimed to establish and characterize an cell model for studying MAmmary Gland mIcrobial Colonization (MAGIC) model. We used the immortalized cell line MCF10A, which expresses the strong polarized phenotype similar to MG ductal epithelium when cultured on a permeable support (Transwell). We analyzed the surface properties of the MAGIC model by gene expression analysis of E-cadherin, tight junction proteins, and mucins and by scanning electron microscopy. To demonstrate the applicability of the model, we tested the adhesion capability of the whole human milk (HM) microbial community and the cellular response of the model when challenged directly with raw HM samples. MCF10A on permeable supports differentiated and formed a tight barrier, by upregulation of CLDN8, MUC1, MUC4, and MUC20 genes. The surface of the model was covered with mucins and morphologically diverse with at least two cell types and two types of microvilli. Cells in the MAGIC model withstood the challenge with heat-treated HM samples and responded differently to the imbalanced HM microbiota by distinctive cytokine response. The microbial profile of the bacteria adhered on the MAGIC model reflected the microbiological profile of the input HM samples. The well-studied MAGIC model could be useful for studies of bacterial attachment to the MG and for studies of biofilm formation and microbiota development.IMPORTANCEThe MAGIC model may be particularly useful for studies of bacterial attachment to the surface of the mammary ducts and for studies of biofilm formation and the development of the human mammary gland (MG) microbiota. The model is also useful for immunological studies of the interaction between bacteria and MG cells. We obtained pioneering information on which of the bacteria present in the raw human milk (HM) were able to attach to the epithelium treated directly with raw HM, as well as on the effects of bacteria on the MG epithelial cells. The MAGIC cell model also offers new opportunities for research in other areas of MG physiology, such as the effects of bioactive milk components on microbial colonization of the MG, mastitis prevention, and studies of probiotic development. Since resident MG bacteria may be an important factor in breast cancer development, the MAGIC tool also offers new opportunities for cancer research.
Topics: Female; Humans; Milk, Human; Mammary Glands, Human; Cytokines; Bacteria; Mucins; Microbiota
PubMed: 38289112
DOI: 10.1128/spectrum.02369-23 -
Avian Pathology : Journal of the W.V.P.A Jun 2024Although anticoccidial drugs have been used to treat avian coccidiosis for nearly a century, resistance, bird harm, and food residues have caused health concerns. Thus,...
Although anticoccidial drugs have been used to treat avian coccidiosis for nearly a century, resistance, bird harm, and food residues have caused health concerns. Thus, was investigated as a possible coccidiosis treatment for broilers. A total of 150 1-day-old male Cobb broiler chicks were treated as follows: G1-Ng: fed a basal diet; G2-Ps: challenged with spp. oocysts and fed basal diet; G3-Clo: challenged and fed basal diet with clopidol; G4-NOa: challenged and fed 0.1% in diet, and G5-NOb: challenged and fed 0.2% . Compared to G2-Ps, in the diet significantly ( < 0.05) decreased dropping scores, lesion scores, and oocyst shedding. Without affecting breast meat colour metrics, improved meat quality characters. At 28 days of age, birds received 0.2% had significantly ( < 0.05) higher serum levels of MDA, T-SOD, HDL, and LDL cholesterol compared to G2-Ps. Serum AST, ALT, and urea levels were all decreased when (0.2%) was used as opposed to G2-Ps. Histopathological alterations and the number of developmental and degenerative stages of spp. in the intestinal epithelium were dramatically reduced by 0.2% compared to G2-Ps. Molecular docking revealed a higher binding affinity of for aldolase, EtAMA1, and EtMIC3, which hindered glucose metabolism, host cell adhesion, and invasion of . Finally, (0.2%) can be used in broiler diets to mitigate the deleterious effects of coccidiosis.
Topics: Animals; Male; Eimeria; Chickens; Molecular Docking Simulation; Coccidiosis; Diet; Oocysts; Meat; Poultry Diseases; Animal Feed; Dietary Supplements
PubMed: 38285881
DOI: 10.1080/03079457.2024.2312133 -
Cell Biochemistry and Function Jan 2024Several cellular processes, including the recovery of misfolded proteins, the folding of polypeptide chains, transit of polypeptides across the membrane, construction...
Several cellular processes, including the recovery of misfolded proteins, the folding of polypeptide chains, transit of polypeptides across the membrane, construction and disassembly of protein complexes, and modulation of protein control, are carried out by DnaJ homolog subfamily A member 1 (DNAJA1), which belongs to the DnaJ heat-shock protein family. It is unknown if DNAJA1 regulates the production of milk in bovine mammary epithelium cells (BMECs). Methionine and leucine increased DNAJA1 expression and nuclear location, as seen by us. In contrast to DNAJA1 knockdown, overexpression of DNAJA1 boosted the production of milk proteins and fats as well as mammalian target of rapamycin (mTOR) and sterol regulatory element binding protein-1c (SREBP-1c). As a result of amino acids, mTOR and SREBP-1c gene expression are stimulated, and DNAJA1 is a positive regulator of BMECs' amino acid-induced controlled milk protein and fat production.
Topics: Animals; Cattle; Amino Acids; Epithelial Cells; Milk Proteins; Sterol Regulatory Element Binding Protein 1; TOR Serine-Threonine Kinases
PubMed: 38269516
DOI: 10.1002/cbf.3918