Catenin Protein Research Articles

Data-based modeling of breast cancer and optimal therapy

Excessive accumulation of β−catenin proteins is a vital driver in the development of breast cancer. Many clinical assessments incorporating immunotherapy with targeted mRNA of β−catenin are costly endeavor. This paper develops novel mathematical models for different treatments by invoking available clinical data to calibrate models, along with the selection and evaluation of therapy strategies in a faster manner with lower cost. Firstly, in order to explore the interactions between cancer cells and the immune system within the tumor microenvironment, we construct different types of breast cancer treatment models based on RNA interference technique and immune checkpoint inhibitors, which have been proved to be an effective combined therapy in pre-clinical trials associated with the inhibition of β−catenin proteins to enhance intrinsic anti-tumor immune response. Secondly, various techniques including MCMC are adopted to estimate multiple parameters and thus simulations in agreement with experimental results sustain the validity of our models. Furthermore, the gradient descent method and particle swarm algorithm are designed to optimize therapy schemes to inhibit the growth of tumor and lower the treatment cost. Considering the mechanisms of drug resistance in vivo, simulations exhibit that therapies are ineffective resulting in cancer relapse in the prolonged time. For this reason, parametric sensitivity analysis sheds light on the choice of new treatments which indicate that, in addition to inhibiting β−catenin proteins and improving self-immunity, the injection of dendritic cells promoting immunity may provide a novel vision for the future of cancer treatment. Overall, our study provides witness of principle from a mathematical perspective to guide clinical trials and the selection of treatment regimens.

Melatonin ameliorates sleep-wake disturbances and autism-like behaviors in the Ctnnd2 knock out mouse model of autism spectrum disorders.

Autism spectrum disorder (ASD) is a prevalent neurodevelopmental disorder characterized by atypical patterns of social interaction and communication, as well as restrictive and repetitive behaviors. In addition, patients with ASD often presents with sleep disturbances. Delta (δ) catenin protein 2 (CTNND2) encodes δ-catenin protein, a neuron-specific catenin implicated in many complex neuropsychiatric diseases. Our previous study demonstrated that the deletion of Ctnnd2 in mice led to autism-like behaviors. However, to our knowledge, no study has investigated the effects of Ctnnd2 deletion on sleep in mice. In this study, we investigated whether the knockout (KO) of exon 2 of the Ctnnd2 gene could induce sleep-wake disorders in mice and identified the effects of oral melatonin (MT) supplementation on Ctnnd2 KO mice. Our results demonstrated that the Ctnnd2 KO mice exhibited ASD-like behaviors and sleep-wake disorders that were partially attenuated by MT supplementation. Overall, our current study is the first to identify that knockdown of Ctnnd2 gene could induce sleep-wake disorders in mice and suggests that treatment of sleep-wake disturbances by MT may benefit to autism-like behaviors causing by Ctnnd2 gene deletion.

A Clinicopathological Study of B-catenin in Colorectal Adenomas

Background and objectives: Colorectal adenomas are considered benign tumors, which originate from the mucus-secreting colonic epithelium creating polyps that protrude into the lumen of the intestine. B-catenin is a member of the catenin protein family and it has a dual function that controls the coordination of cell-cell adhesion and gene transcription. The aim of this study was to investigate the immune expression of B - catenin in colorectal adenomas and to describe B-catenin correlation with clinicopathological parameters. Methods: This study was retrospective in nature and performed on 94 formalin-fixed tissue blocks which were obtained from the histopathology department at Rizgary Hospital and some private laboratories in Erbil city during the period from 2014- 2018, The tissue cuts were prepared in 4 um thick and placed on special salinized slides, samples were deparaffinized and put into a buffer solution for 20 min at 95-99°C, cooled in room temperature for 20 min. Results: The expression of B - catenin was seen in 73.4% of the samples, 41.5% of them with grade II while 31.9% exhibited grade III and 26.6% had grade I B -catenin expression. Except for the type of polyp, there was no statistically significant association between B- catenin expression and the clinicopathological. For patients diagnosed with multiple adenomas, the size of the polyp and grade of expression were remarkably different between the various polyps of the same patient. Conclusions: No statistically significant association between B-catenin expression and clinicopathological parameters except for the type of adenoma.

Clinical significancy of WNT pathway inhibition in various cancers

The tumor microenvironment (TME) plays an important role in the cell cycle. There is a correlation between the Wnt/B–catenin signaling pathway and TME. This article reviews methods of inhibiting Wnt Pathway, a useful process in the treatment of various cancers. Compounds of Wnt/β–Catenin Signaling Pathway, such as TCF–1, have an impact on the differentiation and migration of CD8+ T cells. CCL4 expression is regulated by the beta–catenin protein to recruit CD103+ dendritic cells, which enables CD8+ T cell activation. Inhibition of the Wnt/β–catenin pathway has an impact on ovarian cancer patients’ prognosis, reducing the development of ovarian cancer. Research shows that inhibition of the pathway with the use of the LGK974 inhibitor may boost immunity, especially when applied with a Paclitaxel mix. After treatment, expression of the inhibitory receptors CTLA–4, TIM3, PD–1 on CD8+ T cells decreased. The combination of LGK974 and Paclitaxel can cause the death of tumor cells and significantly inhibit their proliferation. The application of dose–dense Paclitaxel avoids toxicity related to the maximum dose needed to protect the patient's immune system by increasing CD8+ TILs. There are concerns regarding toxicity of the LGK 974, especially for cells dependent on the Wnt/β–catenin pathway to maintain homeostasis. Many Wnt/β–catenin pathway inhibitors are tested against colorectal cancer (CRC) with successful results. These include NSAIDs, porcupine inhibitors, tankyrase inhibitors, Wnt5A inhibitors, and disheveled protein inhibitors.The Wnt/β–catenin pathway, when expressed in Triple Negative Breast Cancer (TNBC), leads to the transition of epithelial to mesenchymal cells. In early clinical development, there are multiple inhibitors (ex. KYA1797K) targeting the Wnt/β–catenin pathway in TNBC cells, which could become a viable anticancer strategy.

A critique on cell signallings involve in colorectal cancer

Colorectal carcinogenesis (CRC) is a significant health issue in developing countries. It ranks as the third most common outcome of cancer death. New drugs are required to lower the prevalence of this ailment despite a multitude of therapeutic choices. As CRC develops, several signaling pathways pathways are activated. Among the important signaling pathways are the p53, Delta-Notch, Wnt/-catenin, Salvador-Warts-Hippo (SWH), and Kelch-like ECH assocd protein 1 pathways. This paper summarises the aetiology of CRC as well as the related death of cells and cell signal transduction pathways.

Regulation effect of miR-7 on intervening colorectal cancer rats with HP infection through Akt/GSK-3β/β-catenin pathway.

This experiment aimed to analysis of the intervention effects of modulating miR-7 on rats with colorectal cancer complicated with HP infection and the effects on (serine/threonine kinase) Akt / (glycogen synthase kinase 3 β） GSK-3 β/ （ β- β - Catenin) β- Influence of the catenin pathway. For this purpose, forty special pathogen-free (SPF) - grade rats of both sexes were randomly divided into 10 control, 10 colorectal cancer and HP infection model groups, 10 up-regulated miR-7, and 10 down-regulated miR-7 groups. Observational analysis of rat colon tissues was performed using the HE staining method. Detection of inflammatory factors [TNF- α、 IL-8, IL-6], detection of miR-7 expression, detection of Akt, GSK-3 using Western blot β、β- Catenin protein expression. Results showed that forty special pathogen-free (SPF) - grade rats of both sexes were randomly divided into 10 control, 10 colorectal cancer and HP infection model groups, 10 up-regulated miR-7, and 10 down-regulated miR-7 groups. Observational analysis of rat colon tissues was performed using the HE staining method. Detection of inflammatory factors [TNF- α、 IL-8, IL-6], detection of miR-7 expression, detection of Akt, GSK-3 using Western blot β、β- Catenin protein expression. It was concluded that modulation of miR-7 in rats with colorectal cancer and HP infection enables regulation of the Akt / GSK-3 β/β- Catenin pathway to improve serum inflammation condition and alleviate HP infection in rats, which played a better role in intervention.

Understanding Molecular Heterogeneity in Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC) is the 5th most common cause of cancer‐related death in the United States with an estimated 32,000 annual deaths. Roughly 30% of HCC cases demonstrate mutations in the CTNNB1 oncogene that encodes a form of the ꞵ‐catenin protein that cannot be degraded. Recently, we developed a mouse model of HCC that genetically represents ~9‐12% of all human HCCs. Here, we combine hydrodynamic tail vein injections with the Sleeping Beauty Transposase – a non‐viral method of expressing genes of interest. Our model uses a clinically observed mutant of ꞵ‐catenin‐(T41A) that is co‐expressed with a mutant form of the nuclear factor erythroid 2–related factor 2 (Nrf2‐G31A) in the livers of wild‐type mice (B+N). This system results in microscopic HCC within 3 weeks and evident macroscopic disease by 7 weeks after induction.ꞵ‐catenin activation can be identified by the expression of its downstream target gene Glutamine Synthetase (GS) in the liver. Patient HCCs with ꞵ‐catenin mutations strongly correlate with GS expression. The evolution and function of GS in b‐catenin‐mutated HCC, remains unclear. Also, there is a gap in the knowledge whether elevated GS expression correlates with HCC biology, which were all subjects of the current study. B+N mouse model of HCC was examined temporally for development of HCC along with GS expression. B+N mice develop HCCs with heterogeneous GS expression in the form of GS‐high and GS‐low to GS‐negative foci. We next asked whether GS expression is associated with cell proliferation or death in the B+N model. We stained tissues for TUNEL and PCNA to mark cell apoptosis and proliferation, respectively in 7‐ and 9.5 weeks post‐injection livers, which were then correlated to GS staining of HCC nodules. While we found no correlation between apoptosis and GS, we found nodules with lower GS expression to have higher proliferation and vice versa. Finally, we asked if eliminating GS expression from B+N HCCs will impact tumor burden. Five weeks after B+N tumor induction in GS‐floxed mice, these animals were injected with AAV8‐TBG‐Cre virus to delete Glul,and mice were harvested after 2‐ or 4.5‐weeks. We found B+N mice with Glul deletion show increased liver weight to body weight ratio, a measurement of HCC burden, compared to controls. The livers of these mice were enriched for GS‐negative tumors. Together, our data suggests that HCC nodules with lower GS expression may outperform GS‐positive foci, and thus are more aggressive compared to the nodules with higher GS expression, which may be more differentiated. The mechanisms of GS heterogeneity and elucidation of its functional significance may help select the most effective targeted therapies for HCC patients with ꞵ‐catenin mutations.

REMOVED: MiR-539-3p inhibits osteoblast activity and bone formation in vivo via targeting LRP-6

X-linked hypophosphatemia (XLH), the most common form of inherited rickets, is characterized by renal phosphate (Pi) wasting, abnormal vitamin D metabolism and defective bone mineralization. It is caused by an inactivation mutation in PHEX gene. Other factors, such as microRNA (miRNA), which regulates biological functions by interfering with specific mRNA translation, also contribute in controlling disease pathogenesis. Recent studies indicate the role of miRNAs in osteoblast functions. The main objective of this study is to identify novel miRNAs, regulated by Phex in osteoblast cells, and to determine whether miR-539-3p, a known tumor suppressor, aids in osteogenesis. MiRNA expression patterns in control and siPhex treated cells were analyzed by miRNA microarray profiling and quantitative RT-PCR. Effect of miR-539-3p on osteoblast differentiation was achieved by transfection with miR-539-3p and anti-miR in mouse calvarial osteoblasts. MiR-539-3p targets were identified by Luciferase reporter gene assay. LRP-6, Wnt3, Beta catenin, p-beta catenin, GSK3β and Lef-1 protein levels were determined by Western blotting. To investigate the function of miR-539-3p in vivo, animal groups were divided into; sham+PBS (ovary intact), ovariectomized (Ovx)+miC, Ovx+miR and Ovx+anti-miR. Chemically modified oligonucleotide miR and anti-miR were injected at 5ug/animal dose via a subcutaneously injection into mice. In PHEX silenced mice calvarial osteoblast cells, miR-539-3p was up-regulated as analyzed by miRNA array profiling and qRT-PCR analysis. Over-expression of miR-539-3p inhibited osteoblast differentiation, whereas inhibition of miR-539-3p function promoted expression of osteoblast-specific genes and alkaline phosphatase (ALP) activity. Target prediction analysis tools and experimental validation by luciferase 3ⲠUTR reporter assay identified LRP-6 as a direct target of miR-539-3p. Over-expression of miR-539-3p led to LRP-6 repression and inhibition of Wnt-Beta catenin signaling. Furthermore, miR-539-3p silencing led to increased bone formation and improved trabecular micro architecture in Ovx mice. Although miR-539-3p is known to be a tumor repressor, we identified a second complementary function of miR-539-3p where it inhibits LRP-6-mediated osteogenesis. Our findings suggest that pharmacological inhibition of miR-539-3p by anti-miR-539-3p could represent a therapeutic strategy for enhancing bone formation in vivo.

Mechanism of dact2 gene inhibiting the occurrence and development of liver fibrosis

There are few reports about the relationship between dact2 and liver fibrosis. The inhibitory mechanism of dact2 in liver fibrosis is not clear, we need to further explore it. In this study has been shown that the dact2 gene can inhibit liver fibrosis. In this experiment, we used lentivirus as a vector to construct a lentivirus vector carrying the dact2 gene and packaged dact2 recombinant lentivirus and its control vector. HSC-T6 infected cells were observed. The effect of dact2 gene expression was activated by Wnt3a HSC-T6 cells. Immunoblot was used to detect α - SMA expression, TGF - β 1, Smad3, Smad7, β - Catenin and CyclinD1. The expression of MMP-2 and TIMP-1 was detected by real-time PCR. At the same time, dact2 recombinant lentivirus was injected into the tail vein. Carbon tetrachloride was used to establish the liver fibrosis model. After 7 weeks of modeling, the staining was used to observe the pathological changes of liver tissue, hydroxyproline was used to analyze the changes of collagen content in liver tissue, the expression of the protein was observed by immunohistochemistry, and the expression of fibrosis-related genes was detected by real-time PCR. Results showed that the dact2 gene expression could inhibit the activation of HSC-T6 cells and reduce the expression of TGF - β 1. The percentage of Smad3, β - Catenin and cyclinD1 protein was 50.02%, 46.73%, 47.58% and 37.50% respectively (P < 0.05).

English

BACKGROUND Colorectal carcinoma is the most frequently encountered malignancy worldwide. The main cause of mortality associated with colorectal malignancy is tumour invasion and metastasis. The major genomic alteration that has been found in colorectal carcinoma is mutation in the adenomatous polyposis gene. Mutated APC causes unrestricted action of the Wnt signalling pathway which results in accumulation of the β - catenin protein in the nucleus responsible for cell proliferation, differentiation and enhanced survival of colorectal epithelial cells. Role of β - catenin expression as a prognostic marker needs to be studied. It will help in aiding the possibility of the future of anti β - catenin targeted chemotherapy for the treatment of colorectal cancers. METHODS A total of 85 samples from histopathologically proven cases of adenocarcinoma colon were taken. Histomorphological features and their immunohistochemical expression of β - catenin were studied. Data thus obtained was analysed using SPSS version 25. Histopathological diagnosis is considered as the gold standard. RESULTS Among the 85 samples of adenocarcinoma 9 case were stage T1 tumours, 24 were stage T2. 31 cases were stage T3, and 21 colorectal cancers were stage T4. Statistically significant correlation was obtained between tumour stage and β - catenin expression. (P value = < 0.001) With regard to tumour grade 24 out of 85 cases were well differentiated and 45 were moderately differentiated, 16 cases were poorly differentiated. Correlation of tumour grading and β - catenin expression also were statistically significant (P value = < 0.001). 45 out of 85 cases were having no nodal metastasis. 22 cases showed metastasis in 1 to 3 lymph nodes. 18 cases were N2. No statistically significant correlation was obtained between lymph node status and β - catenin expression. (P value = 0.003) CONCLUSIONS In this study the membranous, cytoplasmic and nuclear scores of β - catenin shows the linear progression with the advancing stages of colorectal carcinoma. So β - catenin can be used as a prognostic marker in malignant colorectal neoplasm. KEYWORDS Immunohistochemistry, β – Catenin, Colorectal Carcinoma

Abstract LB060: Circulating tumor cells in desmoid tumors

Abstract Introduction: Desmoid tumor (DT) is a rare neoplasm with high local recurrence rates. CTNNB1 mutations are found in most sporadic cases and are located in exon 3, usually in codons 41 or 45 (p.T41A, p.S45F and p.S45P). Circulating tumor cells (CTCs) isolated from the peripheral blood of patients with sarcomas and other neoplasms can be early biomarkers of tumor invasion and dissemination. Moreover, CTCs can also re-colonize their tumors of origin through a process of “tumor self-seeding.” Objectives: To identify CTCs in the peripheral blood of patients with DT; to evaluate their expressions of β- catenin and vimentin proteins. Materials and methods: We conducted a prospective study of patients with initially diagnosed and relapsed DT with measurable disease. Blood samples from each patient were processed and filtered by ISET® (Rarecells, France) for the isolation and quantification of CTCs. The expressions of β -catenin and vimentin in CTCs were analyzed by immunocytochemistry (ICC). Results: A total of 18 patients were included, and all had detectable CTCs. We found a concordance of β-catenin expression in CTCs and in primary tumors in 42.85% of cases by using ICC and immunohistochemistry respectively. Conclusions: Our study identified a high prevalence of CTCs in DT patients. Concordance of β-catenin expression between primary tumor and CTCs bring new perspectives to assess the dynamics of CTCs in the blood microenvironment, opening new avenues for studying the biology and behavior of DT. In addition, these results open the possibility of using CTCs to predict DT dynamics at the time of progression and during treatment. Further studies with larger sample sizes are needed to validate our findings. Citation Format: Alexcia Camila Braun, Fernando Campos, Emne Ali Abdallah, Anna Paula Ruano, Milena Tariki, Fabio Pinto, Celso Abdon de Mello, Vilma Regina Martins, Ludmilla Thome Chinen. Circulating tumor cells in desmoid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB060.

Glucose regulates expression of pro-inflammatory genes, IL-1β and IL-12, through a mechanism involving hexosamine biosynthesis pathway-dependent regulation of α-E catenin.

High glucose levels are associated with changes in macrophage polarisation and evidence indicates that the sustained or even short-term high glucose levels modulate inflammatory responses in macrophages. However, the mechanism by which macrophages can sense the changes in glucose levels are not clearly understood. We find that high glucose levels rapidly increase the α-E catenin protein level in RAW264.7 macrophages. We also find an attenuation of glucose-induced increase in α-E catenin when hexosamine biosynthesis (HB) pathway is inhibited either with glutamine depletion or with the drugs azaserine and tunicamycin. This indicates the involvement of HB pathway in this process. Then, we investigated the potential role of α-E catenin in glucose-induced macrophage polarisation. We find that the reduction in α-E catenin level using siRNA attenuates the glucose-induced changes of both IL-1β and IL-12 mRNA levels under LPS-stimulated condition but does not affect TNF-α expression. Together this indicates that α-E catenin can sense the changes in glucose levels in macrophages via HB pathway and also can modulate the glucose-induced gene expression of inflammatory markers such as IL-1β and IL-12. This identifies a new part of the mechanism by which macrophages are able to respond to changes in glucose levels.

A catenin of the plakophilin-subfamily, Pkp3, responds to canonical-Wnt pathway components and signals

Vertebrate beta-catenin plays a key role as a transducer of canonical-Wnt signals. We earlier reported that, similar to beta-catenin, the cytoplasmic signaling pool of p120-catenin-isoform1 is stabilized in response to canonical-Wnt signals. To obtain a yet broader view of the Wnt-pathway's impact upon catenin proteins, we focused upon plakophilin3 (plakophilin-3; Pkp3) as a representative of the plakophilin-catenin subfamily. Promoting tissue integrity, the plakophilins assist in linking desmosomal cadherins to intermediate filaments at desmosome junctions, and in common with other catenins they perform additional functions including in the nucleus. In this report, we test whether canonical-Wnt pathway components modulate Pkp3 protein levels. We find that in common with beta-catenin and p120-catenin-isoform1, Pkp3 is stabilized in the presence of a Wnt-ligand or a dominant-active form of the LRP6 receptor. Pkp3's levels are conversely lowered upon expressing destruction-complex components such as GSK3β and Axin, and in further likeness to beta-catenin and p120-isoform1, Pkp3 associates with GSK3beta and Axin. Finally, we note that Pkp3-catenin trans-localizes into the nucleus in response to Wnt-ligand and its exogenous expression stimulates an accepted Wnt reporter. These findings fit an expanded model where context-dependent Wnt-signals or pathway components modulate Pkp3-catenin levels. Future studies will be needed to assess potential gene regulatory, cell adhesive, or cytoskeletal effects.

The Small Molecule BC-2059 Inhibits Wingless/Integrated (Wnt)-Dependent Gene Transcription in Cancer through Disruption of the Transducin β-Like 1-β-Catenin Protein Complex.

The central role of β-catenin in the Wnt pathway makes it an attractive therapeutic target for cancers driven by aberrant Wnt signaling. We recently developed a small-molecule inhibitor, BC-2059, that promotes apoptosis by disrupting the β-catenin/transducin β-like 1 (TBL1) complex through an unknown mechanism of action. In this study, we show that BC-2059 directly interacts with high affinity for TBL1 when in complex with β-catenin. We identified two amino acids in a hydrophobic pocket of TBL1 that are required for binding with β-catenin, and computational modeling predicted that BC-2059 interacts at the same hydrophobic pocket. Although this pocket in TBL1 is involved in binding with NCoR/SMRT complex members G Protein Pathway Suppressor 2 (GSP2) and SMRT and p65 NFκB subunit, BC-2059 failed to disrupt the interaction of TBL1 with either NCoR/SMRT or NFκB. Together, our results show that BC-2059 selectively targets TBL1/β-catenin protein complex, suggesting BC-2059 as a therapeutic for tumors with deregulated Wnt signaling pathway. SIGNIFICANCE STATEMENT: This study reports the mechanism of action of a novel Wnt pathway inhibitor, characterizing the selective disruption of the transducin β-like 1/β-catenin protein complex. As Wnt signaling is dysregulated across cancer types, this study suggests BC-2059 has the potential to benefit patients with tumors reliant on this pathway.

Inflammatory and Oxidative Pathways Are New Drug Targets in Multiple Episode Schizophrenia and Leaky Gut, Klebsiella pneumoniae, and C1q Immune Complexes Are Additional Drug Targets in First Episode Schizophrenia.

Breakdown of paracellular and vascular pathways and activated neuroimmune and oxidative pathways was established in (deficit) schizophrenia. The aim of this study was to delineate (a) the differences in these pathways between stable-phase, first (FES) and multiple (MES) episode schizophrenia and (b) the pathways that determine the behavioral-cognitive-physical-psychosocial (BCPS) deterioration in FES/MES. This study included 21 FES and 58 FES patients and 40 healthy controls and measured indicants of serum C1q circulating immune complexes (CIC), leaky gut, immune activation, and oxidative stress toxicity (OSTOX). We constructed a BCPS-worsening index by extracting a latent vector from symptomatic, neurocognitive, and quality of life data. FES was associated with higher IgA CIC-C1q, IgA directed to cadherin, catenin, and plasmalemma vesicle-associated protein, and IgA/IgM to Gram-negative bacteria as compared with FES and controls. In FES patients, the BCPS-worsening score was predicted (48.7%) by IgA to Klebsiella pneumoniae and lowered paraoxonase 1 activity. In MES patients, the BCPS-worsening score was explained (42.7%) by increased tumor necrosis factor-α, OSTOX, and number of episodes. In schizophrenia, 34.0% of the variance in the BCPS-worsening score was explained by IgA to K. pneumoniae, OSTOX, and number of episodes. Increased IgA to K. pneumoniae was the single best predictor of residual psychotic symptoms in FES and MES. This study delineated different mechanistic processes in FES, including breakdown of adherens junctions, bacterial translocation, and IgA CIC-C1q formation, and MES, including immune and oxidative neurotoxic pathways. FES and MES comprise different staging subtypes, i.e., FES and MES with and without worsening.

Effects of latanoprost on the expression of TGF-β1 and Wnt / β-catenin signaling pathway in the choroid of form-deprivation myopia rats

In this study, we investigated the effect of latanoprost on the expression of TGF- β1 and Wnt / β - Catenin signal pathway in the choroid of form-deprivation myopia model rats. Forty rats were randomly divided into two groups: the control group and the FDM model group. Each group had 20 rats. The FDM model group was established by feeding latanoprost daily for 28 days. 15 rats in each group were used to measure the length of the ocular axis and the level of TGF-β1 in choroidal tissue; the remaining 5 rats in each group were used for choroidal fibroblast culture. After modeling, the rats were killed, the length of the ocular axis was measured with a vernier caliper, and the level of TGF - β1 protein and mRNA in the choroidal tissue of each group were measured with RT-PCR method. Results showed that compared with the control group, there was a significant difference in the axial length of the FDM model group (P< 0.05). There was a significant difference in the expression of TGF- β1 protein and mRNA between the two groups (P<0.05). The cultured cells were identified as choroidal fibroblasts by immunocytochemistry. There was no significant difference (P>0.05) in the comparison of GSK3 β protein in choroidal fibroblasts of rats in each group. TGF-β 1 and APC protein in FDM group were significantly lower than those in the control group (P<0.05), while dcl3, p21-gsk3 β and β - Catenin proteins were significantly higher (P<0.05), and there was no significant difference (P>0.05) in the ratio of various indexes protein in FDM + ddk1 group and the comparison of TGF - β1 and APC protein in FDM + ddk1 group and FDM group The expression of dcl3, p21-gsk3 β and β - Catenin decreased significantly (P<0.05). There was no significant difference in the expression of GSK3 β mRNA in the choroidal fibroblasts of each group (P>0.05). The expression of TGF - β 1 and APC mRNA in FDM group was significantly lower than that in the control group (P<0.05), while the expression of dcl3, p21-gsk3 β and β-catenin mRNA in FDM + ddk1 group was significantly higher than that in the control group (P<0.05) >In FDM + ddk1 group, TGF-β 1 and APC mRNA were significantly lower than those in FDM group (P<0.05), while dcl3, p21-gsk3 β and β-Catenin mRNA were significantly higher (P<0.05).

Significant Changes in Trans-Epithelial Barrier Proteins of Adenoid Tissue with Atopic Status in Children.

Adenoid tissue is important in local immune response and epithelial barrier dysfunction of this tissue may contribute to allergies. The aim of this study was to evaluate the relationship between the status of cross-epithelial barrier elements in adenoid tissue lymphoepithelium and inhalant allergen sensitization. Children aged 5-15 years, who underwent adenotonsillectomy, participated in this study. All subjects underwent skin prick testing with environmental inhalant allergens. Occludin, ZO1, e-cadherin, β-catenin, desmoglein, desmoplakin, and connexon-43 were stained immunohistochemically in the adenoid tissues obtained and scored by H-score. We enrolled 76 children, 14 among whom were sensitized to environmental allergens. Among the zonula occludens proteins, median H-scores for occludin, claudin, and ZO-1 were significantly lower in the atopic compared to the nonatopic group respectively (p<0.001). Similarly, median H-scores for e-cadherin and β catenin proteins of the zonula adherens were significantly lower in the atopic group (p<0.001). Both desmoglein and desmoplakin H-scores were significantly lower in the atopic group [60 (50-100) vs 280 (260-300), p<0.001 and 105 (87.5-120) vs 280 (67.25-300), p<0.001 respectively]. Moreover, connexin-43 protein of the gap junction was significantly lower in the atopic group (p<0.001). Adenoid tissue, which is the initial point of contact of inhalant allergens demonstrates epithelial barrier junctional protein, changes in children with inhalant allergen sensitization without clinical allergic disease symptoms. Therefore, it may be concluded that epithelial barrier function plays an important role in the development of allergen sensitization versus tolerance.

Abstract B042: Loss of nuclear alpha catenin contributes to chemoresistance and aggressive disease in African American triple-negative breast cancer patients

Abstract Triple-negative breast cancer (TNBC) is a particularly aggressive, difficult-to-treat subtype of the disease. A well-documented health disparity exists within TNBC: African American (AA) women are more likely to be diagnosed with and die from the disease. Our group reported homozygous deletions in the CTNNA1 gene, which encodes the protein alpha catenin, in AA TNBC. We have undertaken a basic and translational research study to understand the mechanistic role and clinical impact of alpha catenin loss in TNBC, particularly in AA patients. To validate our findings of alpha catenin loss in TNBC, we analyzed over 600 breast cancer patient samples by immunohistochemistry. We found that alpha catenin protein loss was inversely correlated with survival in TNBC. Moreover, loss was observed more frequently in tumors from AA patients. Interestingly, both nuclear and cytosolic alpha catenin adhered to this pattern. While its cytosolic role has been well studied, little is known about nuclear alpha catenin and its role in disease. To understand the nuclear function of alpha catenin, we developed several isogenic cell line pairs. Using CRISPR/Cas9-mediated gene editing, we generated CTNNA1 knockout (KO) BT-549 and MB-MDA-436 cell lines. We also reintroduced CTNNA1 into MDA-MB-468 cells—a line derived from an AA woman with an endogenous deletion in CTNNA1. Using these models, we found that alpha catenin KO cells demonstrated greater capacity for anchorage-independent growth. Loss of alpha catenin also resulted in decreased sensitivity to DNA-damaging chemotherapies including platinum salts (cisplatin, carboplatin), topoisomerase poisons (doxorubicin, etoposide), and the PARP inhibitor olaparib. This is clinically relevant as doxorubicin, carboplatin, and olaparib are approved for treatment of TNBC patients. To identify binding partners, we performed co-immunoprecipitation of alpha catenin from nuclear lysates, followed by mass spectrometry. We found that nuclear alpha catenin can interact directly with ATR, a kinase critical to the DNA damage response (DDR). ATR mediates both the repair of DNA lesions and the G2/M cell cycle checkpoint, which ensures that only cells with undamaged DNA may enter mitosis. We further found that alpha catenin KO cells were more sensitive to inhibitors of ATR, as well as to inhibitors of the G2/M checkpoint proteins Chk1 and Wee1. Tellingly, the KO cells were less sensitive to inhibitors of ATM or DNA-PK, two regulators of alternate DDR pathways. This suggests that nuclear alpha catenin likely plays a specific role in ATR-directed processes. In our study, we have identified nuclear alpha catenin as a tumor suppressor that affects TNBC's susceptibility to chemotherapy by playing a role in the DDR and the G2/M checkpoint. Our data suggest that loss of alpha catenin is more common in AA patients, and is associated with poor prognosis. Therefore, CTNNA1 status may be important in determining appropriate therapeutic strategies in a subset of patients. Citation Format: Rania Bassiouni, Victoria Dirgo, Krystine Mansfield, Ritin Sharma, Lee D. Gibbs, Patrick Pirrotte, Nasreen Vohra, Kevin Gardner, John D. Carpten. Loss of nuclear alpha catenin contributes to chemoresistance and aggressive disease in African American triple-negative breast cancer patients [abstract]. In: Proceedings of the Eleventh AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2018 Nov 2-5; New Orleans, LA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2020;29(6 Suppl):Abstract nr B042.

Galectin-3: A Cardiomyocyte Antiapoptotic Mediator at 24-Hour Post Myocardial Infarction.

Galectin 3 (GAL-3) is a beta galactoside binding lectin that has different roles in normal and pathophysiological conditions. GAL-3 has been associated with heart failure and was linked to increased risk of death in a number of studies. GAL-3 was found to be up regulated in animal models of heart failure as well as myocardial infarction (MI). The objective of his study is to test if high GAL-3 after myocardial infarction has a protective role on the heart through its anti-apoptotic and anti-necrotic functions. Male C57B6/J mice and GAL-3 knockout (KO) mice were used for permanent ligation of the left anterior descending artery of the heart to create infarction in the anterior myocardium. Heart and plasma samples were collected 24 hours after the induction of MI and were used for immunohistochemistry, Tunnel procedure, electron microscopy and enzyme linked immunosorbent assay (ELISA). Our results show that the significant increase in GAL-3 levels in the left ventricle at 24-hour following MI is associated with significant lower levels of pro-apoptotic proteins; cytochrome c, Bax, annexin V, cleaved caspase-3 and a higher levels of anti-apoptotic protein Bcl2 in GAL-3 wild MI group than GAL-3 KO group. We also have identified the anti-apoptotic activity of GAL-3 is mediated through a significant increase in Akt-1, NF kappa-B and beta- catenin proteins. In addition, we have identified the antiapoptotic activity is mediated through a significant lower levels of cathepsin-D protein. We conclude that the increased levels of GAL-3 at 24-hour following MI regulate antiapoptotic mechanisms in the myocardium that will shape the future course of the disease. We also identified that the anti-apoptotic mechanisms are likely mediated through interaction of GAL-3 with Akt-1, NF kappa-B, beta- catenin and cathepsin D proteins.

Investigation of effect of vitamin D receptor, calcium-sensing receptor and β-catenin on cutaneous squamous cell carcinoma

Abstract Background Cutaneous squamous cell carcinoma (cSCC) is a malignant and invasive tumor which is originated from epidermis with a high incidence among non-melanoma skin cancers. The aim of this study was to determine whether vitamin D receptor (VDR), calcium-sensing receptor (CaSR) and beta catenin (β-catenin) proteins have an effect on cSCC. Materials and methods VDR, CaSR and β-catenin proteins in tissue samples of cSCC and control group were analyzed by immunohistochemistry (IHC) and Western blotting (WB) method. IHC findings were statistically evaluated. Results IHC staining density of VDR and β-catenin were higher in cSCC tissue samples than control. The difference between IHC staining density of VDR and β-catenin in the patient and the control groups were statistically significant (p = 0.021, p = 0.021, respectively), but not for CaSR (p = 0.237). While the VDR and β-catenin staining rates obtained by the IHC method could be supported by WB results, the WB bands for CaSR could not be shown. Conclusion The findings suggest that VDR and β-catenin may have an effect on the disease. Further research is required to better understand the role of VDR and β-catenin together on cSCC.