CD44 Positivity Research Articles

#2908 Renal progenitor cells an early non-invasive biomarker of silent kidney injury in Fabry disease

Abstract Background and Aims Fabry's disease (FD), a hereditary condition, is linked to the X chromosome due to a mutation in the gene encoding alpha-galactosidase A (alpha-GalA). This mutation results in a defect in the metabolism of glycosphingolipids, leading to the deposition of globotriaosylceramide (GB3) primarily in organs such as the heart, kidneys, and nervous system. Renal injury can affect any level of the structure, with the podocytes being the most commonly affected due to their low regeneration and high contact with GB3. This leads to proteinuria, and if it exceeds 0.5 g, it is associated with a rapid deterioration of renal function. Renal assessment is carried out through blood analysis, proteinuria measurement, ultrasound, and the gold standard, renal biopsy. However, there is a growing trend towards less invasive biomarkers such as lysoGB3 titration in blood and urine, podocyturia, or renal progenitor cells (RPCs) in renal biopsy material or cell culture. RPCs possess self-renewal, clonogenicity, and multidifferentiation properties, contributing to cellular remodeling. They express specific markers like CD133/CD24 and CD106 in the parietal epithelium of Bowman's capsule. It is known that GB3 expresses CD77. The proliferative response of RPCs can become dysregulated, leading to detachment and elimination in the urine. The aim of the study was to demonstrate the presence of renal progenitor cells (RPCs) in urine as a non-invasive early marker for detecting renal damage in patients with Fabry's disease (FD) and to establish a correlation between RPC presence, GB3 deposition, and their potential association with the degree of renal injury. Method It is an open observational, case-control, single-center study on the urine of patients with Fabry's disease (FD) considered as cases, patients without a history considered as healthy controls, and patients with non-FD renal disease such as Gittelman syndrome or another biopsy-proven cause as positive controls. Renal progenitor cells (RPCs) were isolated using specific markers and flow cytometry, correlating these findings with renal function and albuminuria. Results A total of 75 recent urine samples were processed, corresponding to 59 patients: 16 with Fabry's disease (FD), 11 with Gittelman syndrome, 10 with chronic kidney disease (CKD), and 22 healthy controls. The samples were divided for sediment and microalbuminuria analysis, and separately for the isolation of renal progenitor cells (RPCs). The classification and quantification of the isolated cell types were performed, establishing positivity points using compensation panels with CD3 lymphocyte markers. This was followed by the identification of RPCs through positivity for CD133+/CD24+, CD106+, and CD106– markers. Once RPCs were identified, GB3 marking was carried out using CD77+. Differences were observed among the analyzed patient groups. In healthy controls, there was minimal or no presence of renal progenitor cells (RPCs), except in two patients where isolation was repeated in three different samples, maintaining positivity. Subsequent diagnoses revealed bilateral lithiasis and previously unknown hypertension in these cases. Patients with Gittelman syndrome showed a higher quantity of RPCs compared to healthy controls, but with P > .05. However, clear RPCs positivity was evident in patients with Fabry's disease (FD) and chronic kidney disease (CKD) confirmed by biopsy, with P < .05. Additionally, differences in the number of RPCs were noted based on the degree of renal disease when correlated with the presence of proteinuria < 0.5 g and > 0.5 g, with P < .05. Conclusion With these results, we can conclude that renal progenitor cells (RPCs) may serve as an early biomarker for silent renal injury of various etiologies. Furthermore, if we perform labeling with CD77, we can attribute it to the deposition of GB3 in Fabry's disease (FD).

Abstract 1158: Spatially resolved cell profiling unveils tumor metabolic states associated with immunotherapy response in NSCLC

Abstract Background. Only a subset of non-small cell lung cancer (NSCLC) patients benefit from immune checkpoint inhibitors (ICIs). Thus, there is a clinical need to develop predictive biomarkers for ICIs. Deeper insights from the tumor microenvironment (TME) can lead to identification of spatial features associated with clinical responses to immune checkpoint inhibitors (ICI). In this study, we investigated functional profiles within cell subtypes as a surrogate for ICI clinical endpoints. Methods. We profiled a retrospective cohort of 39 NSCLC tissue cores from 27 patients treated with ICI, using highly multiplexed immunofluorescence (mIF) stained by the Phenocycler Fusion platform (Akoya Biosciences) capturing 45 proteins. Utilizing a deep learning based multiplex imaging analysis pipeline, cells were classified to 15 cell types by known marker expression and were further subclassified by unsupervised clustering. Cells were assigned to the tumor area or TME and more than 1000 spatial features were calculated based on cell type, marker positivity, and area assignments, and were compared between responders and non-responders to ICI therapy using Fisher’s exact and logrank tests. Results. Unsupervised cell subtyping of the 15 cell types identified 43 cell subsets, which were mostly segregated by their metabolism and activation status. Interestingly, all lymphocytes showed a similar pattern of clustering resulting in two clusters of metabolically active and inactive cells. The most significantly differentially expressed proteins between these cell types were oxidative phosphorylation proteins such as CS, SDHA and ATPA5, and other metabolic enzymes such as HK1, GLUT1 and LDHA. We found a direct connection between metabolic state, effector functions and tissue localization as the metabolically active lymphocytes exhibited higher levels of PD-1, MHC class I and II and CD44 positivity, and were more abundant within tumor infiltrating lymphocytes (TILs) and tertiary lymphoid structures (TLS). Unsupervised clustering of tumor cells demonstrated segregation to three main metabolic states - OXPHOS+, OXPHOS- and a third cluster (PPP+) which was characterized by upregulation of ASCT2, a glutamine transporter, as well as pNRF2 and G6PD, regulators of the pentose phosphate pathway. These cells also exhibited higher proliferation rate and CD44, a tumor stemness marker, positivity. All tumors with high content of PPP+ tumor cells (>40%) were resistant to PD-1 blockade (0/8 vs. 10/18 response rate in other tumors, p=0.009) and showed reduced overall survival (OS) rates (median OS of 27.6 vs 90.3 months, p=0.02). Conclusions. Taken together, this study reveals connections between metabolic states, effector functions, and immunotherapy outcome and contributes to the evolving landscape of predictive biomarkers for ICI therapies. Citation Format: James Monkman, Rotem Czertok, Shai Bookstein, Becky Arbiv, Yuval Shachaf, Ron Elran, Kenneth Bloom, Oscar Puig, Ken O'Byrne, Ettai Markovits, Arutha Kulasinghe. Spatially resolved cell profiling unveils tumor metabolic states associated with immunotherapy response in NSCLC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1158.

Frequency of cd44 expression in diagnosed cases of oral Epithelial dysplasia and squamous cell carcinoma.

Oral cavity cancer is one of the most widespread head and neck cancers in the world. 90 percent of oral cavity cancers are squamous cell carcinoma (SCC). Oral Potentially Malignant Disorder (OPMD), are well known to develop into squamous cell carcinoma. CD44 is a glycoprotein present on the cell surface and plays a vital role in cancer cell invasion, migration, metastasis of cancers and prognosis. The present study aims to detect the frequency of CD44 immunomarker expression in epithelial dysplasia and squamous cell carcinoma samples and correlates the expression with histologic grades of this lesion. Cross sectional study at the Department of Pathology, BMSI, JPMC, Karachi between 01-01-2018 to 31-12-2021. 95 patients diagnosed on morphological characterization of the lesion as squamous cell carcinoma/dysplasia were selected by using non-probability purposive sampling technique, among them 41 were diagnosed as epithelial dysplasia and 54 as SCC and were subjected to immunohistochemistry. A total of 95 cases were selected. The mean age was 49.32 years. The majority of cases were seen in 4th and 5th decades. 69 (72.6%) were males and 26 (27.4%) were females with male to female ratio 2.65:1. The closest site was buccal mucosa 75(78.9%). Among 41 cases of dysplasia, 36 showed membranous positivity for CD44 (87.8%), and 45 showed positive membranous CD44 immunoreactivity (83.3%) among 54 cases of squamous cell carcinoma. Increased frequency of CD44 expression was found with increasing grades of dysplasia and squamous cell carcinoma, which may be a possible indicator of malignant transformation and can serve as a prognostic biomarker for oral SCC.

In-situ wound healing by SDF-1-mimic peptide-loaded click crosslinked hyaluronic acid scaffold

Endogenous stem cell-based in-situ tissue regeneration has recently gained considerable attention. In this study, we investigated the potential of a chemokine, SDF-1-mimic peptide (SMP), to promote endogenous stem cell-based in-situ wound healing. Our approach involved the development of a click crosslinked hyaluronic acid scaffold loaded with SMP (Cx-HA + SMP) to release SMP in a wound site. The Cx-HA scaffold maintained its structural integrity throughout the wound healing process and also captured endogenous stem cells. Gradual SMP release from the Cx-HA + SMP scaffold established a concentration gradient at the wound site. In animal wound experiments, Cx-HA + SMP exhibited faster wound contraction compared to Cx-HA + SDF-1. Additionally, Cx-HA + SMP resulted in approximately 1.2–1.6 times higher collagen formation compared to Cx-HA + SDF-1. SMP released from the Cx-HA + SMP scaffold promoted endogenous stem cell migration to the wound site 1.5 times more effectively than Cx-HA + SDF-1. Moreover, compared to Cx-HA + SDF-1, Cx-HA + SMP exhibited higher expression of CXCR4 and CD31, as well as the positive markers CD29 and CD44 for endogenous stem cells. The endogenous stem cells that migrated through Cx-HA + SMP regenerated into wound skin with minimal scar granule formation, similar to the normal tissue. In conclusion, SMP peptide offers greater convenience, while efficiently attracting migrating endogenous stem cells compared to the SDF protein. Our findings suggest that Cx-HA + SMP scaffolds hold promise as a strategy to enhance endogenous stem cell-based in-situ wound healing.

LRRC75A-AS1 Inhibits Chondrogenic Differentiation of Bmscs via Targeting the Mir-140-3p/Wnt/Β-Catenin Pathway.

Bone marrow mesenchymal stem cells (BMSCs) are pluripotent cells with the ability to differentiate into adipocytes, chondrocytes, and osteoblasts. BMSCs are widely used in regenerative medicine and cartilage tissue engineering. Role of lncRNA LRRC75A-AS1 (leucine-rich repeat containing 75A antisense RNA 1) in the chondrogenic differentiation of BMSCs was investigated in this study. BMSCs were isolated from rat bone marrow and then identified using flow cytometry. Alcian blue staining was used to detect chondrogenic differentiation. The effect of LRRC75A-AS1 on chondrogenic differentiation was assessed by western blot. The downstream target of LRRC75A-AS1 was determined by dual luciferase activity assay. BMSCs were identified with positive CD29 and CD44 staining and negative staining of CD34 and CD45. LRRC75A-AS1 was decreased during the chondrogenic differentiation of BMSCs. Silencing of LRRC75A-AS1 increased collagen II (COL II), aggrecan and SOX9 and promoted chondrogenic differentiation. However, over-expression of LRRC75A-AS1 inhibited chondrogenic differentiation. miR- 140-3p was increased during chondrogenic differentiation and interacted with LRRC75A-AS1. miR-140- 3p bind to wnt3a, and inhibition of miR-140-3p up-regulated wnt3a and nuclear β- catenin expression. Wnt3a and nuclear β-catenin were decreased during chondrogenic differentiation. Inhibition of miR-140- 3p attenuated LRRC75A-AS1 deficiency-induced up-regulation of COL II, aggrecan and SOX9. LRRC75A-AS1 suppressed chondrogenic differentiation of BMSCs through down-regulation of miR-140-3p and up-regulation of the wnt/β-catenin pathway.

Immunohistochemistry expression of stem cell markers SOX2, OCT4, CD133 in different grades of meningiomas and correlation with Ki 67 index

BackgroundThere is accumulating evidence of the presence of Cancer Stem Cells (CSCs) within meningiomas (MGs). In the present study, we assessed the Immunohistochemistry (IHC) expression of stem cell markers SOX2, OCT4, CD133 in different grades of MGs and their correlation with Ki 67 index. MethodsA cross-sectional analytical study where 140 MGs cases were graded and classified according to WHO-2021 classification. IHC for SOX2, OCT4, CD133 and Ki-67 was carried out. ResultsOut of 140 cases, 59 cases (42.1%) showed OCT4 positivity, 38 cases (27.1%) showed CD133 positivity and only 21 cases (15%) showed SOX2 positivity. Ki 67 index ranged from 1-30%. Statistically significant difference (Chi Square p-value <0.001) was found between SOX 2, OCT4 and CD133 expressions (positive/negative) and the grades of MG cases. As the grades of MG cases increased, the positive expression of SOX2, OCT 4 and CD 133 also increased. The Mean Ki 67 index value was significantly raised when SOX2, OCT 4 and CD 133 IHC expressions were positive (Student t Test p-value <0.001). ConclusionOur results strongly support the hypothesis of presence of CSCs, in MGs regardless of their grade. We recommend that CSCs IHC markers (SOX2, OCT4 and CD133) to be used on initial diagnosis in all cases of MGs along with Ki-67 index. It can help quantitate CSCs and in turn, recommend closer follow up in patients with high percentage of CSCs to predict early recurrence.

Comparison of human epidermal growth factor receptor 2 and cancer stem cell markers like CD44 and CD133 expressions with clinicopathological parameters in gastric cancer

Objectives: Gastric carcinoma (GC) is the fourth most common cause of cancer-related tumor deaths worldwide. The prognostic significance of CD44, CD133 and human epidermal growth factor receptor 2 (HER2) expression in GC remains controversial. Therefore, we aimed to investigate the relationship of CD44, CD133 and HER2 expression with clinicopathological features in metastatic and non-metastatic GC patients. Methods: A total of 139 patients with GC (68 with metastasis, 71 without metastasis) diagnosed were retrospectively analyzed. CD44 and CD133 expression were determined by immunohistochemical method in all cases. In addition, HER2 overexpression of the tumor was evaluated in patients with metastatic GC. Results: The CD133 positivity rate was 90.6% (n = 126) when all cases were considered, and that for CD44 was 84.9% (n = 118). There was no difference in CD133 and CD44 positivity (intensity or density) rates and between the total scores of metastatic and non-metastatic patients with GC (p &amp;gt; 0.05). HER2 positivity in metastatic cases was detected in 49 (70.1%) patients by immunohistochemical method. No correlation was found between CD133 total score and age, tumor size or depth, and HER2 scores in metastatic or non-metastatic cases (p &amp;gt; 0.05). In the correlation analyzes performed with CD44 scores, only a borderline significant correlation was found between CD44 scores and tumor size (r:0.175; p = 0.047) in non-metastatic cases. Conclusions: We demonstrated associations between CD44/CD133 expression and histological grade in all patients, between CD44 and tumor size in non-metastatic patients, and between HER2 and intestinal type (Lauren) in metastatic patients. The results of this study need to be confirmed by multicenter studies including large case series.

CD44 as a prognostic marker in early colorectal cancer: single center experience “South Egypt cancer institute”, Egypt

ABSTRACT Background Colorectal cancer (CRC) is a common and lethal disease; it remains the third most frequently occurring cancer among both genders. In Egypt, colon cancer represents the ninth most common cancer while rectal cancer ranks the 18th most common cancer by incidence. The application of cancer stem cells (CSC) as prognostic markers in CRC is now studied, among which is CD44, which is implicated in cellular growth, differentiation, survival as well as the metastatic behavior of cancer cells. The prognostic role of CD44 in CRC is still controversial. The study aims at inspecting CD44 immunoreactivity in the epithelium of colorectal cancer specimens and to detect its association with the patients’ clinicopathological features as well as its prognostic significance. Methods This retrospective cross-sectional study included 85 CRC specimens and 16 adenoma specimens collected from the Pathology Department, South Egypt Cancer Institute, during the period from 2018 to 2020. All specimens were stained by anti-CD44 antibody. Results CD44 epithelial expression in colon cancer tissue specimens was substantially higher than in normal mucosa. It was observed that 95% (n = 81) of the studied colorectal carcinoma cases showed positive CD44 epithelial expression compared to distant normal mucosa as controls as well as adenoma cases where only 43.8% (n = 7) of adenoma cases showed positivity with highly significant p-value P = 0.000 and it correlates with cancer progression and aggressiveness. Low H score expression H ≤ 150 had better 3 years disease-free survival DFS estimated by 83.3% compared to 79.6% with high H score expression with significant P-value (P = 0.041). Discussion Our current study showed a statistically significant association between CD44 expression and tumor size (T), lymph node metastasis (N) as well as disease stage with P-values (P = 0.009), (p = 0.000) and (p = 0.000), respectively. CD44 epithelial immunostaining showed to be of an adverse prognostic significance in early CRC associated with more aggressive behavior and worse survival.

Protocol for evaluation and correlation of CD44 immunoexpression in tumor tissue and surgical margins of oral squamous cell carcinoma with three-year survival: A retrospective study

Introduction: The commonest type of cancer in the head and neck region is oral squamous cell carcinoma (OSCC) due to its high rates of occurrence and mortality. Cluster of differentiation 44 (CD44) is a surface glycoprotein present on tumor cells. It regulates cell proliferation, adhesion, migration an invasion of cancer stem cell. Immunoexpression of CD44 in surgical margins indicates poor prognosis of disease and increased risk of local recurrence.[4] Objectives: To evaluate and correlate CD44 immunoexpression in tumor tissue and surgical margins OSCC with three-year survival. Methodology: All tissue sections will be processed for CD44 immunohistochemistry. After that, light microscopic evaluation of CD44 expression will be done. A dark brown color of membranous staining of neoplastic cells will be considered for CD44 positivity; this must be observed in the tumor core, in the basal layer because most stem cells are present in the basal layer of oral mucosa and in the invasive front of tumor. Expected results: The present study will find the immunohistochemical expression of CD44 tumor tissue and surgical margins of OSCC, in order to evaluate the prognosis of OSCC. Immunoexpression may vary in according to their different grades. Conclusions: We hypothesize that, as the disease stage advances, intensity of CD44 expression immunohistochemically increases and it can adversely affect overall survival; hence it is considered as an independent predictor of prognosis of OSCC.

Evaluation of CD44 Expression in Prostatic Adenocarcinoma: An Institutional Study.

Prostate adenocarcinomais the second-most common cause of cancer. Globally, many cancer-related deaths among men were noted due to prostate adenocarcinoma. CD44 plays a key role in mediating cell-to-cell and cell-to-matrix interaction, which further helps to maintain the integrity of tissue and also inhibits tumor metastasis. Cross-sectional study was done on chips from transurethral resections of the prostate (TURP) and prostatic core biopsy specimens. All specimens with clinically diagnosed and histopathologically confirmed prostatic adenocarcinoma were included in the study. Prostatic intraepithelial neoplasia (PIN), recurrent cases, and patients who had undergone radiotherapy/ chemotherapy prior to biopsy were excluded from the study. The sample size for the current study was 57 with an 8% prevalencevalue, 95% confidence interval, and 8% absolute error. Immunoreaction to CD44 antibody is membranous and was evaluated by calculating positively stained cell percentage and stainingintensity. These two parameters were added to obtain a final score; a score of 0-3 was considered as negative, and a score of 4-6 was regarded as positive. A statistically significant difference was only found between Gleason grade (p<0.001), clinical staging (p<0.002), nodal metastasis (p<0.015), and distant metastasis (p<0.020) with CD44 positive expression. The rest of the parameters like PSA (p=0.642) and age (p=0.051) did not correlate with CD44-positive expression. Out of 29 cases with positive CD44 expression, 100% positivity was seen in Gleason's grades 1,2, and 3. This indicates that CD44 expression showed lesser positivity in poorly differentiated carcinoma. CD44 positivity was seen in 83.3% in the T2 stage. An inverse relationship between tumor staging and CD44 expression was observed with positive CD44 expression in lower tumor staging which implies loss of CD44 expression was associated with greater tumor aggressiveness. Lymph node metastasis cases showed more negative CD44 expression (59.5%) and the same was noted in patients without distant metastasis, that is in 61% of the subjects. Conclusion: Cells tend to lose the ability of CD44 expression as they progress from well-differentiated adenocarcinoma to poorly differentiated adenocarcinoma. CD44 expression suggests that the tumor is in a well-differentiated and gland-forming state as compared to Gleason's grade. Loss of CD44 expression suggests tumor aggressiveness. Thus, the upregulation of CD44 expression can be considered as a potential target for targeted therapy. As many targeted and gene therapies are in clinical trials, large-scale multicentered studies are needed for a better understandingof the clinical course of the disease.

#5466 CSF-1R SYSTEM ACTIVATES PARIETAL EPITHELIAL CELLS LEADING TO FOCAL SEGMENTAL GLOMERULOSCLEROSIS (FSGS)

Abstract Background and Aims Focal segmental glomerulosclerosis (FSGS) is the most common glomerular cause leading to end-stage kidney disease. Actual treatment of primary FSGS by immunosuppressive agents presents inconsistent results. Thus, new innovative strategies different from those used to date by taking into consideration podocyte renewal and maintenance or even strategies complementary to immunosuppression are needed. In FSGS, parietal epithelial cells (PECs) switch to an activated phenotype (aPECs) in response to podocyte damage promoting glomerulosclerosis. Colony-stimulating factor (CSF-1) is a hematopoietic growth factor that acts via its specific receptor, the tyrosine kinase receptor CSF-1R. CSF-1 has been detected in sera and renal biopsies from patients with different renal complications, including FSGS, attributing their role and presence to macrophage but not to glomerular cells. The potential cellular and molecular mechanisms involved are also unknown. In this work, we evaluated the implication of CSF-1/CSF-1R axis in the pathogenesis of FSGS and its potential as new pharmacological target point by using specific CSF-1R inhibitors. Specifically, we focused on the modulation of PECS activation by de novo production of CD44 and the preservation of podocyte loss. Method We evaluated the role of the CSF-1/CSF-1R axis as a driver of glomerular damage in FSGS in adriamycin-induced nephropathy (ADR) in mice, the main experimental model to study human FSGS. To this end, we treated or not ADR-animals with CSF-1R specific inhibitors, GW2580 or Ki20227 (n = 5-7 group). We determined the expression and localization of CSF-1R in the glomerulus in tissue by triple immunofluorescence (WT-1, SSeCks and CSF-1R) and their relevance in glomerulosclerosis (PAS and pro-fibrotic genes), the determination of de novo CD44 formation and its correlation with ERK1/2 pathway by immunohistochemistry (IH). We detected podocyte in the glomerulus by WT-1 IH and the localization of aPECS in the glomerular tuff by Claudin-1 and SSeCkS markers. Finally, we used isolated human kidney progenitor cells with and without CSF-1 treatment (n = 6/group) to identify potential key interactors of CSF-1 by RNAseq. We validated genes of interest in the FSGS experimental model. Results We observed a constitutively expression of CSF-1R in the glomerulus of control mice that significantly increased in ADR-treated mice, specifically in podocytes (WT1) and PECs (SSeCks), confirmed with mRNA expression. ADR-treated mice showed important events of sclerotic glomerulus and pro-fibrotic genes as collagen that was significantly reversed by the treatment with CSF-1R inhibitors (p&amp;lt;0.001). Results reflected a CSF-1R inhibitor-dependent renal function recovery with the use of the inhibitors with a reduction of proteinuria and an increase of glomerular filtration rate (p&amp;lt;0.001). We found a positive CD44 staining both in Bowman's capsule and inner the tuff of ADR-treated mice, accompanied by an increase of ERK1/2 activation. CSF-1R inhibitors significantly reduced the percentage of glomeruli with de novo CD44 production. Remarkably, podocyte depletion was also preserved with very similar levels to non-treated animals (p&amp;lt;0.001). For RNAseq results, 227 differentially expressed genes (considering a criterion of a probability of differential expression &amp;gt;0.9 and a |M| &amp;gt; 1) were subjected to enrichment analysis. The top significant gene ontology terms were mainly involved with interferon-induced genes. Genes involved in this pathway were validated in the FSGS model, showing an increase in mice treated with ADR compared to controls and alleviated with CSF-1R inhibitors (p&amp;lt;0.001). Conclusion In this study we propose a novel therapeutic strategy to FSGS-associated pathology based on the inhibition of CSF1-R activity having an impact on reducing aPECs, glomerulosclerosis, proteinuria, improving renal function and preserving the podocyte-progenitor phenotype, thus, in podocyte preservation against damage.

Cancer stem cell isolation and characterization from lung cancer cells

Lung cancer is one of the most common cancers in the world and a leading cause of death. Cancer stem cells (CSCs) are the cells responsible for tumor initiation. CSCs features include self-renewal capacity, differentiation, high invasion-migration, and resistance to chemotherapy. CSCs are characterized by specific surface markers. CD133 is widely used as a marker of CSCs in many cancers, including lung cancer. CD133-positive cells are an important marker of lung cancer stem cells because they show characteristics of lung cancer CSCs. CSCs are less sensitive and/or resistant to conventional treatments, remaining viable and regenerating tumors after treatment. Treatments that can target CSCs will be able to destroy CSCs more effectively, leading to the disappearance of tumors. Currently, attempts are being made to develop therapeutics that can effectively target CSCs. These studies concern the isolation and characterization of CSCs. In this study, CD133-positive CSCs and CD133-negative cells were isolated from H460 cells, a large cell lung cancer model, by MACS method. CSCs properties were investigated by tumorSphere formation assay (2% agar plate), hanging drop assay and qPCR method. CD133 positivity of H460 cells was determined as 0.3%. It has been observed that CD133-positive cells show CSCs-specific self-renewal properties. It was determined that the CD133 gene expression difference between CD133-positive CSCs and CD133-negative cells was approximately 25-fold. It has been observed that CD133-positive CSCs express more pluripotency genes (such as OCT-4, SOX-2 and KLF-4) compared to CD133-negative cells.

Proteinuria in thrombotic microangiopathy is associated with partial podocytopathy

ABSTRACT Background. Thrombotic microangiopathy (TMA) results in acute kidney injury, but the cause of heavy proteinuria in this disorder is puzzling. The goal of this study was to determine if there were significant effacement of foot processes and CD133-positive hyperplastic podocytes in TMA to explain the proteinuria. Methods. The study included 12 negative controls (renal parenchyma removed from renal cell carcinoma) and 28 thrombotic microangiopathy due to different etiologies. The percent of foot process effacement was estimated, and proteinuria level was obtained for each TMA case. Both groups of cases were stained for CD133 by immunohistochemical method, and the number of positive CD133 in hyperplastic podocytes was counted and analyzed. Results. Nineteen (19) of 28 (68%) TMA cases had nephrotic range proteinuria (urine protein/creatinine >3). Twenty-one (21) of 28 (75%) TMA cases showed positive CD133 staining in scattered hyperplastic podocytes within Bowman’s space but was absent in control cases. The percent of foot process effacement (56 ± 4%) correlated with proteinuria (protein/creatinine ratio 4.4 ± 0.6) (r = 0.46, p = .0237) in TMA group. Conclusion. Our data indicate that the proteinuria in TMA can be associated with significant effacement of foot processes. CD133-positive hyperplastic podocytes can be seen in the majority of TMA cases of this cohort, indicating a partial podocytopathy.

Triple-negative mouse breast cancer initiating cells show high expression of beta1 integrin and increased malignant features.

Triple-negative breast cancer (TNBC) is a subtype of breast cancer that exhibits aggressive tumor phenotypes, including rapid metastasis and tumor recurrence. Integrins belong to the family of transmembrane glycoproteins involved in regulating cell adhesion, proliferation, and differentiation through cell-cell and cell-extracellular matrix interactions. Aberrant β1 integrin signaling has been implicated in cancer invasion and metastasis processes. The present work aimed to investigate the role of β1 integrin in TNBC cancer progression using a mouse 4T1 cell line as a model system. We have sorted a subset of tumor-initiating cells (TICs) from the 4T1 cell line based on CD133 positivity by flow cytometry. RT-PCR and protein analysis studies showed the transcriptional upregulation of β1 integrin and its downstream target focal adhesion kinase in 4T1-TICs compared to parental 4T1 cells. In addition, the expression of β1 receptors in TICs is significantly higher than in parental population cells. Furthermore, in vitro cellular assays revealed that CD133+ TICs have higher clonogenic ability, invasion, and sphere formation potential. These findings suggest that β1 integrin has a potential role in TNBC invasion and metastasis. Hence, β1 integrin could be a possible factor for future targeted cancer therapies.

Expression of circulating tumour cells in oral squamous cell carcinoma: An ex vivo pilot study.

Despite advances in diagnostics and therapeutics, two-thirds of oral cancer patients present with advanced disease, which increases both the morbidity and mortality risk. Circulating tumour cells (CTCs) are released in the circulation by primary tumours and have been demonstrated to have significant correlations between their occurrence and disease progression. To characterize the circulating tumour cells in subjects with histologically diagnosed oral squamous cell carcinoma (OSCC). This pilot study was undertaken with ten fresh blood samples (6 ml each). Five samples from apparently healthy individuals and five OSCC samples were cultured and subjected to flow cytometric analysis for CD44 expression. Immunostaining was done using CD44 and EpCAM markers. Several cells in OSCC samples showed EpCAM and CD44 positivity following immunostaining. However, flow cytometry performed with CD44 alone was not specific for OSCC samples. Hence, proving that CD44 and EpCAM when used in conjunction can help to characterize CTCs. The findings of our study suggest that the demonstration of CTCs is feasible and helps in understanding of disease progression and metastatic risk. Sensitive detection of CTCs from blood samples can serve as an implicit tool in early cancer diagnosis and prognosis through liquid biopsy which in itself is minimally invasive and time-saving.

Glomerular parietal epithelial expression of CD44 in minimal change nephrotic syndrome and primary focal segmental glomerulosclerosis: A clinico-pathological study.

Minimal change nephrotic syndrome (MCNS) and focal segmental glomerulosclerosis (FSGS) are the two common causes of nephrotic syndrome (NS) in both children and adults with overlapping clinical features, but with distinct prognostic and therapeutic implications. The distinction between these relies entirely on histopathology, which can sometimes be difficult. CD44 is expressed by activated parietal epithelial cells, plays a role in matrix deposition and thus in the pathogenesis of FSGS. To assess the expression of CD44 in MCNS and FSGS and to evaluate its association with the known clinical and histopathological prognostic factors. Thirty cases each of MCNS and FSGS were studied. The clinical, laboratory, histopathological, and CD 44 immunohistochemical data were recorded. The findings were analyzed and correlated. A P value of < 0.05 was considered statistically significant. Statistical association was noted between CD44 positivity and serum creatinine (p = 0.031), estimated glomerular filtration rate (p = 0.040), segmental sclerosis (p < 0.001), tubular atrophy (p = 0.027), interstitial fibrosis (p = 0.027), and histological diagnosis (p < 0.001). The sensitivity, specificity, positive predictive, and negative predictive values were 90%, 76.67%, 79.41% and 88.46%, respectively. CD44 immunostain can effectively distinguish MCNS from FSGS. The congruent results of CD44 positivity with known prognostic factors support the possibility of using the CD44 marker as a predictive tool in selecting high-risk patients and offering appropriate therapeutic measures.

Possible role of ALDH1 and CD44 in lip carcinogenesis.

Lip squamous cell carcinoma (LSCC) accounts for 12% of all head and neck cancers. It is caused by chronic exposure to ultraviolet light solar radiation and related to previous actinic cheilitis (AC). This study aimed to investigate the immunostaining of the putative cancer stem cells (CSC) markers ALDH1 and CD44 in AC (n=30) and LSCC (n=20). ALDH1 positivity was found to be statistically higher in LSCC than in AC lesions (p=0.0045), whilst CD44 expression was statistically higher in AC than in LSCC lesions (p=0.0155). ALDH1+ cells in AC lesions were associated with specific clinical features: a younger age (<60 years old), the female gender, white skin, not smoking or consuming alcohol, and a fast evolution, and not associated with the chronic exposure to UV radiation (p<0.0001). CD44 positivity was associated with patients who were male, feoderm, smoked, consumed alcohol, underwent occupational exposure to UV-radiation, and demonstrated lesions with log-time evolution (p<0.0001). ALDH1 + cells were associated with mild dysplasia using a system from the World Health Organization (WHO), and with a low risk of malignant transformation, according to the binary system (p<0.0001). CD44+ cells were also associated with moderated dysplasia, according to the WHO system. In LSCC, ALDH1 + cells were positively associated with patients who were older (≥ 60 years old), smokers, and with those who consumed alcohol (p<0.0001). CD44 + cells in LSCC were associated with older (≥ 60 years old) patients as well, but also with female patients, white skin, non-smokers, and individuals who did not consume alcohol (p<0.0001), all of whom showed distinct patterns in pre- and malignant lesions of both markers. Additionally, in LSCC, both ALDH1 and CD44 staining were associated with smaller tumor sizes (T1/T2; p<0.0001). In summary, although both ALDH1 and CD44 were associated with the presence of dysplasia in AC lesions, the present findings suggest that ALDH1 and CD44 may be activated by different etiopathogenic pathways, predominantly in distinct steps of oral carcinogenesis. CD44 would thus be more significantly related to the potentially malignant lesion, while ALDH1 would be closely linked to malignancy.

The Potential Role of CD44 and CD133 in Colorectal Stem Cell Cancer

Colorectal cancer (CRC) is the most preventable malignancy globally, with a high mortality rate. Cancer stem cells (CSCs) are found previously in multiple types of cancer; CRC is one of them, and it has been correlated with several biomarkers. The two most essential markers related to colorectal CSCsare CD44 and CD133, which play a significant role in diagnosis, treatment, and prognosis. Unfortunately, the CSCs with positive CD44 and CD133 biomarkersillustrated an alarming prognosis. Several trials were trying to target those markers to improve the prognosis and cure. We aimed to review the papers that relate to the two markers in terms of diagnosis, treatment, and prognosis.

CD133 is an independent predictive and prognostic marker in metastatic breast cancer

CD133 is a transmembrane glycoprotein and is considered the most common cell surface marker to identify cancer stem cells in hematological and solid tumors, including breast cancer. To evaluate the impact of immunohistochemical expression of CD133 on response rate and survival in metastatic breast cancer, as well as to correlate it with various demographics and clinicopathological characteristics. One-hundred metastatic breast cancer patients were prospectively recruited at the Medical Oncology Department at South Egypt Cancer Institute during the period from January 2018 to January 2020. There was a statistically significant correlation between CD133 positive patients with various adverse clinicopathological parameters such as high grade (p= 0.013), higher tumor (p= 0.001), and nodal staging (p= 0.024) during a median follow-up time of 17 months. In addition, Cases with CD133 positive expression had a significantly lower survival time than those with negative expression (3-years OS 37.4% versus 85.5%, p= 0.024). Regarding the response rate, CD133 positive patients had a lower response rate than negative patients (50% versus 54%, p= 0.012). Positive CD133 is correlated with poor prognosis in metastatic breast cancer patients.

Aldehyde dehydrogenase 1 and CD44 serve as prognostic markers in patients with breast cancer

Background and Objective: Breast cancer is the second leading cause of cancer-related death in women globally, and its prevalence is rising quickly, particularly in low- and middle-income nations. Despite significant advancements in treatment options, a small percentage of individuals with advanced-stage breast cancer have a dismal prognosis. The most extensively utilised markers for identifying breast cancer stem cells are ALDH1 and CD44 (BCSCs). The goal of this study was to look into the expression of ALDH1 and CD44 in breast carcinoma and see if there was any correlation with other clinicopathological factors to see if they might be used to predict prognosis in patients with breast cancer. Methods: This study comprised 30 women with breast cancer who were undergoing mastectomy. Immunohistochemistry (IHC) labelling with an ALDH1, CD44 primary antibody was used to assess ALDH1, CD44 levels in paraffin-embedded tissues. The percentage of positive cells was used to assess the expression level, which was then associated with clinicopathological characteristics. Results: Out of 30 patients, 23 (76%) had CD44 positive; out of 30 patients, 21 had CD44 positivity (70 percent). ALDH1 expression was linked to the number of lymph nodes, while CD44 expression was linked to tumour size. Conclusions: In breast cancer, ALDH1 and CD44 expression acts as an independent prognostic indicators. However, bigger population-based prospective patient trials are needed to confirm these findings.