Dharmendra Kumar Maurya

Naive human umbilical cord matrix derived stem cells significantly attenuate growth of human breast cancer cells in vitro and in vivo.

The effect of un-engineered (naïve) human umbilical cord matrix stem cells (hUCMSC) on the metastatic growth of MDA 231 xenografts in SCID mouse lung was examined. Three weekly IV injections of 5x10(5) hUCMSC significantly attenuated MDA 231 tumor growth as compared to the saline-injected control. IV injected hUCMSC were detected only within tumors or in close proximity to the tumors. This in vivo result was corroborated by multiple in vitro studies such as colony assay in soft agar and [(3)H]-thymidine uptake. These results suggest that naïve hUCMSC may be a useful tool for cancer cytotherapy.

Human umbilical cord matrix-derived stem cells expressing interferon-β gene significantly attenuate bronchioloalveolar carcinoma xenografts in SCID mice

Mesenchymal stem cells derived from the human umbilical cord matrix (hUCMSCs) have great potential for therapeutic use for multiple diseases. The strategy that uses therapeutic gene-transfected hUCMSCs as cellular vehicles for targeted biologic agent delivery has solved the problem of short half-life or excessive toxicity of biological agent(s) in vivo. Interferon-beta (IFN-beta) has demonstrated a potent antitumor effect on many types of cancer cell lines in vivo. The aim of this study was to determine the anti-cancer effect of IFN-beta gene-transfected hUCMSCs (IFN-beta-hUCMSCs) on cells derived from bronchioloalveolar carcinoma, a subset of lung adenocarcinoma that is difficult to treat. The co-culture of a small number of IFN-beta-hUCMSCs with the human bronchioloalveolar carcinoma cell lines H358 or SW1573 significantly inhibited growth of both types of carcinoma cell lines. The culture medium conditioned by these cells also significantly attenuated the growth of both carcinoma cells, but this attenuation was abolished by adding anti-IFN-beta antibody. Finally, systemic administration of IFN-beta-hUCMSCs through the tail vein markedly attenuated growth of orthotopic H358 bronchioloalveolar carcinoma xenografts in SCID mice by increasing apoptosis. These results clearly indicate that IFN-beta-hUCMSCs caused cell death of bronchioloalveolar carcinoma cells through IFN-beta production, thereby attenuating tumor growth in vivo. These results indicate that IFN-beta-hUCMSCs are a powerful anti-cancer cytotherapeutic tool for bronchioloalveolar carcinoma.

Angiotensin II type 2 receptor signaling significantly attenuates growth of murine pancreatic carcinoma grafts in syngeneic mice

BackgroundPancreatic cancer is one of the most aggressive human malignancies, with a very poor prognosis. To evaluate the effect of angiotensin II (Ang II) type 2 receptor (AT2) expression in the hosts body on the growth of pancreatic carcinoma, we have investigated the growth of mouse pancreatic ductal carcinoma grafts in syngeneic wild type and AT2 receptor-deficient (AT2-KO) mice.MethodsThe role of AT2 receptor-signaling in stromal cells on the growth of murine pancreatic carcinoma cells (PAN02) was studied using various in vitro and in vivo assays. In vivo cell proliferation, apoptosis, and vasculature in tumors were monitored by Ki-67 immunostaining, TUNEL assay, and von Willebrand factor immunostaining, respectively. In the co-culture study, cell proliferation was measured by MTT cell viability assay. All the data were analyzed using t-test and data were treated as significant when p < 0.05.ResultsOur results show that the growth of subcutaneously transplanted syngeneic xenografts of PAN02 cells, mouse pancreatic ductal carcinoma cells derived from the C57/BL6 strain, was significantly faster in AT2-KO mice compared to control wild type mice. Immunohistochemical analysis of tumor tissue revealed significantly more Ki-67 positive cells in xenografts grown in AT2-KO mice than in wild type mice. The index of apoptosis is slightly higher in wild type mice than in AT2-KO mice as evaluated by TUNEL assay. Tumor vasculature number was significantly higher in AT2-KO mice than in wild type mice. In vitro co-culture studies revealed that the growth of PAN02 cells was significantly decreased when grown with AT2 receptor gene transfected wild type and AT2-KO mouse-derived fibroblasts. Faster tumor growth in AT2-KO mice may be associated with higher VEGF production in stromal cells.ConclusionsThese results suggest that Ang II regulates the growth of pancreatic carcinoma cells through modulating functions of host stromal cells; Moreover, Ang II AT2 receptor signaling is a negative regulator in the growth of pancreatic carcinoma cells. These findings indicate that the AT2 receptor in stromal fibroblasts is a potentially important target for chemotherapy for pancreatic cancer.

Over-expression of angiotensin II type 2 receptor gene induces cell death in lung adenocarcinoma cells.

The endogenous angiotensin II (Ang II) type 2 receptor (AT2) has been shown to mediate apoptosis in cardiovascular tissues. Thus, the aim of this study was to explore the anti-cancer effect of AT2 over-expression on lung adenocarcinoma cells in vitro using adenoviral (Ad), FuGENE, and nanoparticle vectors. All three gene transfection methods efficiently transfected AT2 cDNA into lung cancer cells but caused minimal gene transfection in normal lung epithelial cells. Ad-AT2 significantly attenuated multiple human lung cancer cell growth (A549 and H358) as compared to the control viral vector, Ad-LacZ, when cell viability was examined by direct cell count. Examination of annexin V by flow cytometry revealed the activation of the apoptotic pathway via AT2 over-expression. Western Blot analysis confirmed the activation of caspase-3. Similarly, poly (lactide-co-glycolic acid) (PLGA) biodegradable nanoparticles encapsulated AT2 plasmid DNA were shown to be effectively taken up into the lung cancer cell. Nanoparticle-based AT2 gene transfection markedly increased AT2 expression and resultant cell death in A549 cells. These results indicate that AT2 over-expression effectively attenuates growth of lung adenocarcinoma cells through intrinsic apoptosis. Our results also suggest that PLGA nanoparticles can be used as an efficient gene delivery vector for lung adenocarcinoma targeted therapy.

Therapy with un-engineered naïve rat umbilical cord matrix stem cells markedly inhibits growth of murine lung adenocarcinoma

BackgroundLung cancer remains the leading cause of cancer-related mortality despite continuous efforts to find effective treatments. Data from the American Cancer Society indicate that while the overall incidence of lung cancer is declining, it continues to rise in women. Stem cell-based therapy has been an emerging strategy to treat various diseases. The purpose of this paper is to determine the efficacy of an intrinsic anti-cancer effect of rat umbilical cord matrix stem cells (UCMSCs) on lung cancer.MethodsA mouse syngeneic lung carcinoma model was used to test the basic ability of UCMSCs to control the growth of lung cancer. Lung tumors were experimentally induced by tail vein administration of Lewis lung carcinoma (LLC) cells derived from the lung of C57BL/6 mouse. Rat UCMSCs were then administered intratracheally five days later or intravenously on days 5 and 7. The tumor burdens were determined by measuring lung weight three weeks after the treatment.ResultsCo-culture of rat UCMSCs with LLC significantly attenuated the proliferation of LLC cells as monitored by MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide), a tetrazole cell proliferation assay, thymidine uptake, and direct cell counts. In vitro colony assays with rat UCMSCs as feeder layers markedly reduced LLC colony size and number. Co-culture of rat UCMSCs with LLCs causes G0/G1 arrest of cancer cells. This is evident in the decrease of cyclin A and CDK2 expression. The in vivo studies showed that rat UCMSC treatment significantly decreased tumor weight and the total tumor mass. Histological study revealed that intratracheally or systemically administered rat UCMSCs homed to tumor areas and survived for at least 3 weeks without any evidence of differentiation or adverse effects.ConclusionsThese results indicate that rat UCMSCs alone remarkably attenuate the growth of lung carcinoma cells in vitro and in a mouse syngeneic lung carcinoma graft model and could be used for targeted cytotherapy for lung cancer.

Angiotensin II bi-directionally regulates cyclooxygenase-2 expression in intestinal epithelial cells

We previously demonstrated that angiotensin II (Ang II) receptor signaling is involved in azoxymethane-induced mouse colon tumorigenesis. In order to clarify the role of Ang II in COX-2 expression in the intestinal epithelium, the receptor subtype-specific effect on COX-2 expression in a rat intestinal epithelial cell line (RIE-1) has been investigated. Ang II dose- and time-dependently increased the expression of COX-2, but not COX-1 mRNA and protein. This stimulation was completely blocked by the AT1 receptor antagonist but not the AT2 receptor antagonist. Ang II and lipopolysaccharide (LPS) additively induced COX-2 protein in RIE-1 cells, whereas the LPS-induced COX-2 expression was significantly attenuated by low concentrations of Ang II or the AT2 agonistic peptide CGP-42112A only in AT2 over-expressed cells. These data indicate that Ang II bi-directionally regulates COX-2 expression via both AT1 and AT2 receptors. Control of COX-2 expression through Ang II signaling may have significance in cytokine-induced COX-2 induction and colon tumorigenesis.

Non-random tissue distribution of human naïve umbilical cord matrix stem cells

AIM To determine the tissue and temporal distribution of human umbilical cord matrix stem (hUCMS) cells in severe combined immunodeficiency (SCID) mice. METHODS For studying the localization of hUCMS cells, tritiated thymidine-labeled hUCMS cells were injected in SCID mice and tissue distribution was quantitatively determined using a liquid scintillation counter at days 1, 3, 7 and 14. Furthermore, an immunofluorescence detection technique was employed in which anti-human mitochondrial antibody was used to identify hUCMS cells in mouse tissues. In order to visualize the distribution of transplanted hUCMS cells in H&E stained tissue sections, India Black ink 4415 was used to label the hUCMS cells. RESULTS When tritiated thymidine-labeled hUCMS cells were injected systemically (iv) in female SCID mice, the lung was the major site of accumulation at 24 h after transplantation. With time, the cells migrated to other tissues, and on day three, the spleen, stomach, and small and large intestines were the major accumulation sites. On day seven, a relatively large amount of radioactivity was detected in the adrenal gland, uterus, spleen, lung, and digestive tract. In addition, labeled cells had crossed the blood brain barrier by day 1. CONCLUSION These results indicate that peripherally injected hUCMS cells distribute quantitatively in a tissue-specific manner throughout the body.

Abstract 3328: Cytotherapy with naïve rat umbilical cord matrix stem cells significantly attenuates growth of murine pancreatic cancer cells and increases mouse survival in syngeneic mice

Pancreatic cancer is one of the most aggressive human malignancies, with a very poor prognosis. It is the fourth leading cause of cancer-related morbidity and mortality in the United States. MSC derived from the human umbilical cord matrix (UCMSC) have a great potential for therapeutic use in cancer. UCMSC-based targeted gene/therapeutic-delivery and successful tumor attenuation has been demonstrated. However, the targeted stem cell therapy would be significantly safer and more applicable to human patients if unmodified postnatal stem cells alone were shown to cause tumor cell death and regress tumors, without any foreign gene transfection. Accordingly, the aim of this study was to determine the anti-cancer effect of un-engineered naive rat UCMSC on the growth of murine pancreatic ductal carcinoma in vitro and in vivo. A mouse peritoneal model was used to test the ability of un-engineered rat UCMSC to control growth of pancreatic cancer. In addition, MTT, direct cell count, [ 3 H] thymidine uptake, and soft agar colony assays were carried out as in vitro studies. Co-culture of rat UCMSC with PAN02 murine pancreatic carcinoma cells (UCMSC:PAN02, 1:6 and 1:3) caused G0/G1 arrest and significantly attenuated the proliferation of PAN02 carcinoma cells as monitored by MTT assay, direct cell counts, and [ 3 H] thymidine uptake assay. Rat UCMSCs also significantly reduced PAN02 colony size and number as measured by soft agar colony assay. The in vivo mouse studies showed that rat UCMSC treatment significantly decreased the peritoneal PAN02 tumor burden 3 weeks after tumor transplantation and increased mouse survival time. Histological study revealed that intraperitoneally administered rat UCMSC survived for at least 3 weeks and the majority were found near or inside the tumor. These results indicate that naive rat UCMSC alone remarkably attenuate the growth of pancreatic carcinoma cells in vitro and in a mouse peritoneal model. Thus, these studies imply that UCMSC could be a potential tool for targeted cytotherapy for pancreatic cancer. This work was supported by Kansas State University (KSU) Terry C. Johnson Center for Basic Cancer Research, KSU College of Veterinary Medicine Dean9s fund, Kansas State Legislative Appropriation, NIH P20RR017686, P20RR015563 and R21 CA135599. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3328.