Role Of Bone Marrow-derived Cells Research Articles

A novel role for bone marrow-derived cells to recover damaged keratinocytes from radiation-induced injury

Exposure to moderate doses of ionizing radiation (IR), which is sufficient for causing skin injury, can occur during radiation therapy as well as in radiation accidents. Radiation-induced skin injury occasionally recovers, although its underlying mechanism remains unclear. Moderate-dose IR is frequently utilized for bone marrow transplantation in mice; therefore, this mouse model can help understand the mechanism. We had previously reported that bone marrow-derived cells (BMDCs) migrate to the epidermis-dermis junction in response to IR, although their role remains unknown. Here, we investigated the role of BMDCs in radiation-induced skin injury in BMT mice and observed that BMDCs contributed to skin recovery after IR-induced barrier dysfunction. One of the important mechanisms involved the action of CCL17 secreted by BMDCs on irradiated basal cells, leading to accelerated proliferation and recovery of apoptosis caused by IR. Our findings suggest that BMDCs are key players in IR-induced skin injury recovery.

The role of bone marrow-derived cells in venous thromboembolism.

Venous thrombosis is a life-threatening condition with high morbidity and mortality. Abnormal functioning of different cells in the blood is an integral part of its pathogenesis. In this review, we describe the contribution of bone marrow-derived cells to the development of this debilitating disease. We present both epidemiological and clinical data demonstrating involvement of various cell types in venous thrombosis, and discuss potential mechanisms underlying these effects. Modern concepts including recently discovered new paradigms in thrombosis, such as neutrophil extracellular traps, mast cells, and polyphosphate, are summarized.

The role of bone marrow-derived cells in the origin of liver cancer revealed by single-cell sequencing.

Objective: Epithelial cancers often originate from progenitor cells, while the origin of hepatocellular carcinoma (HCC) is still controversial. HCC, one of the deadliest cancers, is closely linked with liver injuries and chronic inflammation, which trigger massive infiltration of bone marrow-derived cells (BMDCs) during liver repair.Methods: To address the possible roles of BMDCs in HCC origination, we established a diethylnitrosamine (DEN)-induced HCC model in bone marrow transplanted mice. Immunohistochemistry and frozen tissue immunofluorescence were used to verify DEN-induced HCC in the pathology of the disease. The cellular origin of DEN-induced HCC was further studied by single cell sequencing, single-cell nested PCR, and immunofluorescence-fluorescence in situ hybridization.Results: Studies by using single cell sequencing and biochemical analysis revealed that HCC cells in these mice were coming from donor mice BMDCs, and not from recipient mice. Furthermore, the copy numbers of mouse orthologs of several HCC-related genes previously reported in human HCC were also altered in our mouse model. DEN-induced HCCs exhibited a similar histological phenotype and genomic profile as human HCCs.Conclusions: These results suggested that BMDCs are an important origin of HCC, which provide important clues to HCC prevention, detection, and treatments.

Differentiation and roles of bone marrow-derived cells on the tumor microenvironment of oral squamous cell carcinoma.

The stroma affects the properties and dynamics of the tumor. Previous studies have demonstrated that bone marrow-derived cells (BMDCs) possess the capability of differentiating into stromal cells. However, the characteristics and roles of BMDCs in oral squamous cell carcinoma remain unclear. The current study therefore investigated their locations and features by tracing green fluorescent protein (GFP)-labeled BMDCs in a transplantation mouse model. After irradiation, BALB-c nu-nu mice were injected with bone marrow cells from C57BL/6-BALB-C-nu/nu-GFP transgenic mice. These recipient mice were then injected subcutaneously in the head with human squamous cell carcinoma-2 cells. Immunohistochemistry for GFP, Vimentin, CD11b, CD31 and α-smooth muscle actin (SMA), and double-fluorescent immunohistochemistry for GFP-Vimentin, GFP-CD11b, GFP-CD31 and GFP-α-SMA was subsequently performed. Many round-shaped GFP-positive cells were observed in the cancer stroma, which indicated that BMDCs served a predominant role in tumorigenesis. Vimentin(+) GFP(+) cells may also be a member of the cancer-associated stroma, originating from bone marrow. Round or spindle-shaped CD11b(+) GFP(+) cells identified in the present study may be macrophages derived from bone marrow. CD31(+)GFP(+) cells exhibited a high tendency towards bone marrow-derived angioblasts. The results also indicated that spindle-shaped α-SMA(+) GFP(+) cells were not likely to represent bone marrow-derived cancer-associated fibroblasts. BMDCs gathering within the tumor microenvironment exhibited multilineage potency and participated in several important processes, such as tumorigenesis, tumor invasion and angiogenesis.

Bone-marrow-derived cell-released extracellular vesicle miR-92a regulates hepatic pre-metastatic niche in lung cancer.

Metastatic tumors have been shown to establish a supportive pre-metastatic niche (PMN) in distant organs, which in turn determines disseminated tumor cells' targeting of such organs. PMN is formed through the recruitment of bone-marrow-derived cells (BMDCs); however, the role of BMDCs in PMN formation is not fully understood. On the basis of RNA-seq data and bioinformatic analysis, secretion of extracellular vesicle (EV) miR-92a by BMDCs of lung cancer-bearing mice contributes to the establishment of liver PMN. Both BMDC-derived EVs and miR-92a mimics potentiate the activation of hepatic stellate cells (HSCs), subsequently increasing extracellular matrix (ECM) deposition in mice. Consequently, remodeling of the liver microenvironment enhanced immunosuppressive cell accumulation and cancer cell attachment. EVs miR-92a directly suppressed its target SMAD7, leading to the enhancement of transforming growth factor-β signaling in HSC. Elevated levels of circulating miR-92a are found in the sera of lung cancer patients, and EVs isolated from these patients have a similar ability to increase HSCs activation and ECM protein expression. Our study reveals the sequential steps of liver PMN formation in lung cancer, providing critical mediators that prepare PMN in the liver, and identifies new targets that offer valuable options for diagnosis and therapeutic intervention.

Differentiation of bone marrow-derived cells toward thermogenic adipocytes in white adipose tissue induced by the β3 adrenergic stimulation.

Bone marrow provides progenitors of several types of cells, including muscle and white adipocytes, ensuring peripheral tissue homeostasis. However, the role of bone marrow-derived cells (BMCs) in induction of thermogenic adipocytes is unresolved. The purpose of this study is to examine whether BMCs are involved in the emergence of thermogenic adipocytes through adrenergic activation. Irradiation of mice with 8 Gy of X-ray-depleted BMCs and peripheral blood mononucleated cells (PBMCs), which in turn impaired induction of uncoupling protein 1 (UCP1) through administration of β3 adrenergic receptor agonist, CL 316,243 (CL), in inguinal white adipose tissue (iWAT). In contrast, CL-induced UCP1 induction in brown adipose tissue was unaffected by BMC depletion. Transplantation of normal BMCs into mice depleted of BMCs recovered PBMC levels and rescued the ability of iWAT browning by CL. Furthermore, analyses of mice transplanted with green fluorescent protein (GFP)-labeled BMCs revealed that the number of GFP-positive BMCs and PBMCs were significantly decreased by CL and that GFP-positive stromal cells and GFP-positive UCP1-expressing multilocular adipocytes appeared in iWAT after CL administration, demonstrating differentiation of BMC-derived preadipocytes into UCP1-expressing thermogenic adipocytes. These results unveiled a crucial role of the BMC as a nonresident origin for a subset of thermogenic adipocytes, contributing to browning of white adipose tissue.-Yoneshiro, T., Shin, W., Machida, K., Fukano, K., Tsubota, A., Chen, Y., Yasui, H., Inanami, O., Okamatsu-Ogura, Y., Kimura, K. Differentiation of bone marrow-derived cells toward thermogenic adipocytes in white adipose tissue induced by the β3 adrenergic stimulation.

A role for bone marrow-derived cells in diabetic nephropathy.

Diabetes mellitus causes systemic disorders. We previously demonstrated that diabetic condition forced bone marrow-derived cells (BMDCs) to express TNF-α, leading to the development of diabetic neuropathy in mice. Here, we hypothesized that these abnormal BMDCs are also involved in diabetic nephropathy. To test our hypothesis, mice were irradiated to receive total bone marrow (BM) from the transgenic mice expressing green fluorescent protein before diabetes was induced by streptozotocin. Confocal microscopy showed that the diabetic glomerulus had more BMDCs compared with the nondiabetic glomerulus. Most of these cells exhibited endothelial phenotypes, being negative for several markers, including podocin (a maker of podocyte), α8 integrin (mesangial cell), CD68, and F4/80 (macrophage). Next, the total BM of diabetic mice was transplanted into nondiabetic mice to examine if diabetic BM per se could cause glomerular injury. The recipient mice exhibiting normal glycemia developed albuminuria and mesangial expansion with an increase in capillary area. The number of BMDCs increased in the glomerulus of the recipient mice. These cells were found to exhibit the endothelial phenotype and to express TNF-α. These data suggest that diabetic BMDCs per se could initiate glomerular disease. Finally, eNOS knockout mice were used to examine if residential endothelial injury could attract BMDCs into the glomerulus. However, endothelial dysfunction due to eNOS deficiency failed to attract BMDCs into the glomerulus. In summary, BMDCs may be involved in the development of diabetic nephropathy.-Nobuta, H., Katagi, M., Kume, S., Terashima, T., Araki, S., Maegawa, H., Kojima, H., Nakagawa, T. A role for bone marrow-derived cells in diabetic nephropathy.

Estrogen enhances tumor growth and angiogenesis indirectly via mediation of bone marrow‑derived cells as well as directly through stimulation of tumor and endothelial cells.

Estradiol (E2) is a prime culprit for enhancing the progression of female hormone‑related cancers. Bone marrow‑derived cells (BMDCs) have been found to play a pivotal role in tumor growth. Estrogen receptors (ERs) are also found on certain subtypes of BMDCs, in addition to endothelial cells (ECs) and certain tumor cells. However, the role of BMDCs in E2‑induced tumor biology is still unclear. Thus, the effects of E2 on ER‑negative 4T1 breast cancer growth, the mobilization and recruitment of BMDCs, and interactions among BMDCs, ECs, and 4T1 cells were investigated. The results showed that E2 potentiated 4T1 tumor growth and angiogenesis in mice subjected to sham operation, ovariectomy (OVX), or OVX and E2 replacement treatment. E2 supplementation in mice with OVX upregulated the transcription of stromal cell‑derived factor‑1 (SDF‑1) mRNA in tumor tissues and enhanced the recruitment of BMDCs into tumor tissues invivo. E2 deficiency significantly decreased proangiogenic CXCR4+, β3+, Sca‑1+ and CXCR4+β3+ BMDCs circulating in the peripheral blood. Cell‑based system analyses showed that E2 augmented the transcription of β3 mRNA in ECs, increased the adhesion of BMDCs to ECs. In addition, E2 enhanced the BMDC‑induced EC proliferation and migration, the BMDC‑induced 4T1 proliferation and the 4T1‑stimulated EC proliferation in addition to enhancing the proliferation of tumor cells and the migration of ECs invitro. Therefore, E2 enhanced the growth of breast tumors by stimulating tumor cells and ECs directly, as well as by increasing proangiogenic BMDC mobilization and recruitment leading to augmentation of the tumor and EC functions indirectly by cell proliferation assay. These findings reveal a separate mechanism via which E2 promotes the growth of female hormone‑dependent tumors, which may be useful in explorations of new therapies for related cancers.

The Role of Bone Marrow-Derived Cells during Ectopic Bone Formation of Mouse Femoral Muscle in GFP Mouse Bone Marrow Transplantation Model.

Multipotential ability of bone marrow-derived cells has been clarified, and their involvement in repair and maintenance of various tissues has been reported. However, the role of bone marrow-derived cells in osteogenesis remains unknown. In the present study, bone marrow-derived cells during ectopic bone formation of mouse femoral muscle were traced using a GFP bone marrow transplantation model. Bone marrow cells from C57BL/6-Tg (CAG-EGFP) mice were transplanted into C57BL/6 J wild type mice. After transplantation, insoluble bone matrix (IBM) was implanted into mouse muscle. Ectopic bone formation was histologically assessed at postoperative days 7, 14, and 28. Immunohistochemistry for GFP single staining and GFP-osteocalcin double staining was then performed. Bone marrow transplantation successfully replaced hematopoietic cells with GFP-positive donor cells. Immunohistochemical analyses revealed that osteoblasts and osteocytes involved in ectopic bone formation were GFP-negative, whereas osteoclasts and hematopoietic cells involved in bone formation were GFP-positive. These results indicate that bone marrow-derived cells might not differentiate into osteoblasts. Thus, the main role of bone marrow-derived cells in ectopic osteogenesis may not be to induce bone regeneration by differentiation into osteoblasts, but rather to contribute to microenvironment formation for bone formation by differentiating tissue stem cells into osteoblasts.

Abstract 1628: Early events in metastatic spread: new approaches using targeted therapeutics to disrupt formation of the pre-metastatic niche and development of lung metastases

Abstract Solid tumors release soluble factors causing migration of a subset of normal bone marrow derived cells (BMDCs) - primarily VEGFR-1+ hematopoietic progenitor cells and VEGFR-2+ circulating endothelial progenitor cells - from the bone marrow to organ sites. These normal BMDCs self-assemble into a pre-metastatic niche- a micro-environment eventually hosting migratory tumor cells from the primary site. Disruption of this niche or selective targeting the migratory tumor cells may inhibit metastatic spread. The significance and specific characteristics of the BMDCs is still unclear. We have previously observed migration of VEGFR-1+ BMDCs to the lung and lymph nodes, and VEGFR-2+ BMDCs to liver, lung and lymph nodes. Using GFP+ BMDC’s from genetically engineered mice, we characterized the role that recruitment of BMDCs may play in breast cancer metastasis. We assessed the BMDC recruitment profile in lethally irradiated female nu/nu mice transplanted with GFP+ BMDCs from donor mice, followed by orthotopic placement of MDA-MB-231/luc cells or injected with MDA-MB-231/luc conditioned media for 30 days. Flow cytometry results show a gradual increase in the recruitment of CD11b+VEGFR-1+ cells in all the tissues examined from tumor-bearing mice. Recruitment of these cells to the liver in mice treated only with MDA-MB-231/Luc conditioned media was also apparent, but the increase was not as high as in livers of mice with the orthotopically-placed tumors. Recruitment of CD11b+VEGFR-2+ BMDCs was also observed but only in liver and lung. Interestingly, conditioned media seemed to recruit this subset of cells more strongly to these two tissues than signaling from orthotopic tumors. Spleen and lymph node showed minimal recruitment of CD11b+VEGFR-2+ BMDCs. We have previously developed the GrB/VEGF fusion construct- a novel pro-apoptotic fusion protein which specifically targets cells harboring VEGFR2 and have utilized this agent to specifically target BMDCs which are VEGFR2 +. Our preliminary data indicate that treatment with GrB/VEGF does not significantly alter the recruitment of VEGFR-1+ or VEFGR-2+ cells to lungs, when assessed two weeks after the final treatment. We observed increased recruitment of F4/80+ macrophages to the lung. On the other hand, CD11b+Gr-1- BMDCs were significantly reduced following GrB/VEGF treatment, although the F4/80+ subpopulation carrying this signature increased. Studies are ongoing to determine whether systemic administration of this agent can disrupt the formation of the niche and the eventual establishment of metastatic tumors. Understanding the role of BMDCs in metastatic spread and the formation of the pre-metastatic niche and their role in the early development of metastasis will be critical in designing targeted therapeutic approaches to inhibit the metastatic process. Research sponsored, in part, by the Clayton Foundation for Research. Citation Format: Khalid A. Mohamedali, Lawrence H. Cheung, Michael G. Rosenblum. Early events in metastatic spread: new approaches using targeted therapeutics to disrupt formation of the pre-metastatic niche and development of lung metastases [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1628. doi:10.1158/1538-7445.AM2017-1628

Abstract A59: A rodent in situ glioma allograft model to study bone marrow-derived cell migration

Abstract Radiotherapy (RT) induced hypoxia recruits bone marrow-derived cells (BMDCs) to tumors through the upregulation of Stromal Derived Factor-1, a chemokine that binds to receptor CXCR4 present on BMDCs. The infiltration of BMDCs, which includes endothelial progenitors, pericyte progenitors, tumor-associated macrophages, immature monocytes, VEGFR+ hemangiocytes, and CD11b+ myeloid cells has been implicated in supporting tumor establishment by contributing to tumor growth, angiogenesis, vasculogenesis, inflammation, and immune suppression. In this work, we have used a rodent glioma in situ allograft model to study the recruitment/migration of BMDCs post RT. Female albino C57 (8-10 weeks old, Jackson Laboratories) animals underwent +DsRed bone marrow transplant (B6.Cg-Tg(CAG-DsRed*MST)1Nagy/J donor mouse). Following engraftment, glioma cells were cranially implanted with 1x105 glioma cells (GL261-Luc, Perkin Elmer), at an average size of 26.4mm3 indicated by a bioluminescence signal of 5.28x108 photons/sec/cm2/sr, animals underwent 6Gy (RadSource 2000, 250kVp, 25mA) of irradiation limited to a 1cm wide slit centered on the tumor. Bioluminescence (tumor size) and fluorescence imaging (+DsRed BMDCs) was performed prior to and at 1- and 2-weeks post-RT to assess tumor growth and measure the influx of +DsRed BMDCs. In addition, flow cytometry was used to measure the number of +DsRed BMDCs in the tumor and brain. To study the effect of BMDCs in tumor regression, tumors were irradiated at a smaller size (9.3mm3) with 10Gy irradiation. At 1-week post-RT, flow cytometry was used to measure the number of BMDCs (CD45+) in the tumor. In our model 6Gy radiation was able to stall the growth of the tumor. Irradiated tumors did not significantly change in tumor size at either 1- or 2-weeks post irradiation compared to pre-RT size. In addition, irradiated tumors were significantly smaller than untreated tumors (p=0.0008) 1-week post-RT. Using flow cytometry 1-week post irradiation, we observed a significant increase in the number of +DsRed BMDCs in the irradiated tumor (23.5% of cells +DsRed BMDCs) compared to the irradiated brain (6.17% cells +DsRed BMDCs, p=0.03) as well as a significant increase in the tumors (16.6% cells +DsRed BMDCs) compared to the brain (11.1% cells +DsRed BMDCs p=0.02). There was no difference in the number of +DsRed BMDCS in tumors ± irradiation or brains ± irradiation. We found similar results in our tumor regression model using flow cytometry 1-week post-RT; we observed an increase in the number of BMDCs CD45+ in the tumor (~20% of cells CD45+) compared to control tumors (~6% of cells CD45+). Indicating that tumor size at the onset of RT did not effect the migration of BMDCs. Fluorescence imaging did not measure any significant change in the DsRed signal at 1- or 2-weeks post-irradiation. We have successfully shown that post-RT there is an increase in BMDCs migration at 1-week post RT regardless of tumor size at RT. Further experimentation is needed to better elucidate the role of BMDCs in tumor inflammation and immune suppression as well as angiogenesis and vasculogenesis. Our next step is to perform immunohistochemistry on our collected tumor tissue to map out the spatial location of migrated BMDCs within the tumor. Citation Format: Janice A. Zawaski, Tien T. Tang, M Waleed Gaber. A rodent in situ glioma allograft model to study bone marrow-derived cell migration. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2016 Oct 20-23; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2017;5(3 Suppl):Abstract nr A59.

Context-dependent effects of SOCS3 in angiotensin II-induced vascular dysfunction and hypertension in mice: mechanisms and role of bone marrow-derived cells.

Carotid artery disease is a major contributor to stroke and cognitive deficits. Angiotensin II (Ang II) promotes vascular dysfunction and disease through mechanisms that include the IL-6/STAT3 pathway. Here, we investigated the importance of suppressor of cytokine signaling 3 (SOCS3) in models of Ang II-induced vascular dysfunction. We examined direct effects of Ang II on carotid arteries from SOCS3-deficient (SOCS3(+/-)) mice and wild-type (WT) littermates using organ culture and then tested endothelial function with acetylcholine (ACh). A low concentration of Ang II (1 nmol/l) did not affect ACh-induced vasodilation in WT but reduced that of SOCS3(+/-) mice by ∼50% (P < 0.05). In relation to mechanisms, effects of Ang II in SOCS3(+/-) mice were prevented by inhibitors of STAT3, IL-6, NF-κB, or superoxide. Systemic Ang II (1.4 mg/kg per day for 14 days) also reduced vasodilation to ACh in WT. Surprisingly, SOCS3 deficiency prevented most of the endothelial dysfunction. To examine potential underlying mechanisms, we performed bone marrow transplantation. WT mice reconstituted with SOCS3(+/-) bone marrow were protected from Ang II-induced endothelial dysfunction, whereas reconstitution of SOCS3(+/-) mice with WT bone marrow exacerbated Ang II-induced effects. The SOCS3 genotype of bone marrow-derived cells did not influence direct effects of Ang II on vascular function. These data provide new mechanistic insight into the influence of SOCS3 on the vasculature, including divergent effects depending on the source of Ang II. Bone marrow-derived cells deficient in SOCS3 protect against systemic Ang II-induced vascular dysfunction.

Bone marrow-derived cells in ocular neovascularization: contribution and mechanisms

Ocular neovascularization often leads to severe vision loss. The role of bone marrow-derived cells (BMCs) in the development of ocular neovascularization, and its significance, is increasingly being recognized. In this review, we discuss their contribution and the potential mechanisms that mediate the effect of BMCs on the progression of ocular neovascularization. The sequence of events by which BMCs participate in ocular neovascularization can be roughly divided into four phases, i.e., mobilization, migration, adhesion and differentiation. This process is delicately regulated and liable to be affected by multiple factors. Cytokines such as vascular endothelial growth factor, granulocyte colony-stimulating factor and erythropoietin are involved in the mobilization of BMCs. Studies have also demonstrated a key role of cytokines such as stromal cell-derived factor-1, tumor necrosis factor-α, as well as vascular endothelial growth factor, in regulating the migration of BMCs. The adhesion of BMCs is mainly regulated by vascular cell adhesion molecule-1, intercellular adhesion molecule-1 and vascular endothelial cadherin. However, the mechanisms regulating the differentiation of BMCs are largely unknown at present. In addition, BMCs secrete cytokines that interact with the microenvironment of ocular neovascularization; their contribution to ocular neovascularization, especially choroidal neovascularization, can be aggravated by several risk factors. An extensive regulatory network is thought to modulate the role of BMCs in the development of ocular neovascularization. A comprehensive understanding of the involved mechanisms will help in the development of novel therapeutic strategies related to BMCs. In this review, we have limited the discussion to the recent progress in this field, especially the research conducted at our laboratory.

Malignant Transformation in Glioma Steered by an Angiogenic Switch: Defining a Role for Bone Marrow-Derived Cells.

Low-grade gliomas, such as pilocytic astrocytoma and subependymoma, are often characterized as benign tumors due to their relative circumscription radiologically and typically non-aggressive biologic behavior. In contrast, low-grades that are by their nature diffusely infiltrative, such as diffuse astrocytomas and oligodendrogliomas, have the potential to transform into malignant high-grade counterparts and, given sufficient time, invariably do so. These high-grade gliomas carry very poor prognoses and are largely incurable, warranting a closer look at what causes this adverse transition. A key characteristic that distinguishes low- and high-grade gliomas is neovascularization: it is absent in low-grade gliomas, but prolific in high-grade gliomas, providing the tumor with ample blood supply for exponential growth. It has been well described in the literature that bone marrow-derived cells (BMDCs) may contribute to the angiogenic switch that is responsible for malignant transformation of low-grade gliomas. In this review, we will summarize the current literature on BMDCs and their known contribution to angiogenesis-associated tumor growth in gliomas.

Abstract 2282: K-RasV12 bone marrow cell reconstitution causes lung cancer in wild type mice

Abstract The potential of recruited bone-marrow-derived cells (BMDCs) to act as a source of malignancy was disputed for a long time. A role for BMDCs in tissue repair has also been proposed, since these cells are capable of homing to the lung and differentiating into multiple epithelial cell lineages so thus contributing in lung tissue repair and regeneration. We tested the ability of bone marrow cells containing an inducible oncogene to initiate lung adenocarcinoma. Since the role of the K-Ras gene in lung cancer is well known, for our experimental strategy, we used a mouse model expressing oncogenic K-RasV12 (K-Ras+/V12RERTert/ert). This model generates a specific neoplastic phenotype induced by the K-RasV12 mutation. Mice usually develop lung tumors with complete penetrance evolving from hyperplasia, through adenoma, to adenocarcinoma, when the oncogene expression is induced by 4-hydroxitamoxifen (4-OHT) administration. Following lethal irradiation, wild-type mice were transplanted with donor bone marrow cells derived from K-RasV12 mice and, once the bone marrow was reconstituted, treated with 4-OHT. Mice were monitored and sacrificed at subsequent post-induction time points (0, 3.5, 5.5, 8.5, 11, 13.5, 15, and 16 months) and lungs and peripheral blood were collected. Analysis of the H&amp;E-stained lung sections from both reconstituted K-RasV12 and control mice revealed diffuse hyperplasia, possibly due to irradiation. Epithelial lesions, adenomas and/or adenocarcinomas, were detected in 4 out of 8 K-RasV12-reconstituted animals at 3.5, 5.5, 13.5 and 15 months post oncogene induction while no lesions were detected in the control group (transplanted but not induced mice). In addition, the K-Ras implication on the origin of the tumor was confirmed by X-Gal staining of lung lesions in reconstituted mice. In our model, -geo is indeed a marker protein coexpressed along with the same K-RasV12 transcript by a bicistronic strategy. Our results suggest a direct implication of BMDCs in lung epithelial tumor development and emphasize the need of further studies to dissect the mechanisms regulating BMDCs’ recruitment to injured tissues and their role in tumor initiation and/or progression. Citation Format: Elena Belloni, Ines Martin-Padura, Elvira Gerbino, Stefania Orecchioni, Fulvia Fusar-Imperatore, Paola Marighetti, Giovanni Bertalot, Pier Giuseppe Pelicci, Francesco Bertolini. K-RasV12 bone marrow cell reconstitution causes lung cancer in wild type mice. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2282. doi:10.1158/1538-7445.AM2015-2282

Evidence of cell fusion in carcinogen-induced mice gastric carcinoma.

The role of bone marrow-derived cells in gastric cancer formation was not fully understood. In this study, bone marrow from female green fluorescent protein transgenic mice was transplanted into male wild-type mice to generate sex-mismatched chimeric mice. The chimeric mice were treated with carcinogen to induce gastric cancer. At time of sacrifice, 18.2 % (2/11) of mice showed severe dysplasia and 25 % (3/12) of mice successfully induced with cancer. Fluorescence in situ hybridization results showed that bone marrow-derived cells participated in renewal of gastric mucosa and cell fusion was observed in both precancerous lesions and adenocarcinoma, but no sign of fusion was observed in squamous cell carcinoma. Our findings suggest that bone marrow-derived cells participate in renewal of gastric mucosa during chronic damage and might have acquired the phenotype of gastric epithelial cells through cell fusion. Fusion between gastric epithelial cells and bone marrow-derived cells was involved in increased carcinogenesis.

Abstract 610: Context-Dependent Effects of Suppressor of Cytokine Signaling 3 (SOCS3) in Angiotensin II-Induced Vascular Dysfunction: Mechanisms and Role of Bone Marrow-Derived Cells

Angiotensin II (Ang II) promotes vascular disease and hypertension, in part, by activating the interleukin-6 (IL-6)/signal transducer and activator of transcription 3 (STAT3) pathway. Although SOCS3 regulates this pathway in the immune system, its role in vascular disease and hypertension is unknown. In this study, we investigated the role of SOCS3 in a model of Ang II-induced vascular disease. To exam direct effects, carotid arteries from wild-type (WT) and SOCS3 haplodeficient (SOCS3 +/- ) mice were incubated with Ang II for 22 hrs, followed by examination of endothelial function using acetylcholine (Ach). Relaxation to Ach was similar in all arteries incubated with vehicle. A low concentration of Ang II (1 nmol/L) did not affect Ach-induced vasodilation in WT mice, but reduced that of SOCS3 +/- mice by ~50% (P&lt;0.05). Ang II-induced impairment was prevented by inhibitors of STAT3, IL-6, NF-κB, or a scavenger of superoxide. Responses to nitroprusside were similar in all groups. We also tested the impact of SOCS3 in vivo by systemically infusing Ang II (1.4 mg/kg per day) for 14 days via osmotic mini-pumps. Ach-induced vasodilation in carotid and resistance arteries in brain from WT mice was reduced by ~60% (P&lt;0.05). Surprisingly, deficiency in SOCS3 prevented the majority of Ang II-induced endothelial dysfunction without affecting the pressor response to Ang II. Lethally irradiated WT mice reconstituted with SOCS3 +/- bone marrow were protected from Ang II-induced endothelial dysfunction (P&lt;0.05), while reconstitution of irradiated SOCS3 +/- mice with WT bone marrow exacerbated Ang II-induced vascular dysfunction (P&lt;0.05). WT into WT and SOCS3 +/- into SOCS3 +/- bone marrow chimeras exhibited vascular function consistent with non-irradiated controls. The pressor response to Ang II was reduced by ~50% in WT mice reconstituted with bone marrow from SOCS3 +/- mice (P&lt;0.05). These data suggest SOCS3 exerts divergent local versus systemic effects on Ang II-induced vascular dysfunction. In the face of SOCS3 deficiency, bone marrow-derived cells protect against Ang II-induced vascular dysfunction and hypertension.

Tumor Dormancy and the Angiogenic Switch: Possible Implications of Bone Marrow- Derived Cells

Although escape from tumor dormancy has long been recognized as an important problem in the treatment of cancer, the molecular and cellular regulators underlying this transition remain poorly understood. The inability of the cancer cells to induce a complete and successful process of angiogenesis can result in tumor dormancy. In this case, the acquisition of sufficient angiogenic potential will result in the escape from indolence and in the initiation of tumor mass expansion. This stage in disease progression is known as the angiogenic switch. It is now becoming clear that the induction of the angiogenic switch is controlled by dynamic and complex biological processes involving the cancer cells, the associated stromal microenvironment and distant normal host cells, mostly from the bone marrow. Indeed, intricate tumor-host interactions are increasingly recognized as critical features of cancer. In particular, infiltrating cells of the immune system are crucial constituents of tumors and an important source of the growth stimulatory signals to the tumor cells. Tumor cells are surrounded by stromal cells, such as fibroblasts, lymphocytes, neutrophils, macrophages and mast cells, which communicate via a complex network of intercellular signaling pathways, mediated by surface adhesion molecules, cytokines and their receptors. However, the possible roles of these cells and molecules in the maintenance of micro-tumors in an occult state and in the induction of exit from the dormant state are not fully elucidated. In this review, we summarize recent findings and the current understanding of the role of bone marrow-derived cells, their recruitment into tumors and their interactive crosstalk with tumor cells, in leading to either the maintenance of, or exit from, tumor dormancy. Understanding the mechanisms of tumor growth and metastatic recurrence after periods of indolence is crucial for improving early detection, as well as increasing the cure rate for cancer patients.

Increased numbers of bone marrow-derived cells in parathyroid adenoma.

In primary hyperparathyroidism (PHPT), the increased levels of parathyroid hormone (PTH) result in mobilisation of bone-marrow-derived cells (BMCs) into peripheral blood. However, the fate of these cells is still unknown. In this study, we sought to investigate cells with typical surface markers of BMCs within parathyroid adenomas (PA) of patients with primary hyperparathyroidism. We therefore investigated PA and normal parathyroid glands (NPG) of 15 patients with PHPT by immunohistochemistry and PCR. mRNA levels for CD31, CD34 and CD45 were significantly increased in PA compared to NPG. Immunohistochemical staining for CD31 and CD34 revealed a significantly higher vessel density in PA. Furthermore, scattered single cells expressing CD31, CD34 or CD45 were significantly augmented compared to normal parathyroid glands and directly correlated with vessel density. mRNA levels of SDF-1 was increased whereas its major inhibitor dipeptidylpeptidase IV (DPP IV) is decreased in PA, suggesting that the SDF-1 axis plays a role in the migration of BMCs into PA. These data indicate a possible role of BMCs in the pathophysiology of PA of patients with PHPT.

Helicobacter pylori infection and stem cells at the origin of gastric cancer.

Helicobacter pylori infection is now recognized as the main and specific infectious cause of cancer in the world. It is responsible for gastric adenocarcinomas of both intestinal and diffuse types, which are the long-term consequences of the chronic infection of the gastric mucosa. Case-control studies have shown an association between the two, recognized as early as 1994 and further substantiated by interventional studies in which H. pylori eradication has led to the prevention of at least part of the gastric cancers. Experimental studies have highlighted the role of bone marrow-derived cells (BMDCs) and particularly mesenchymal stem cells, in the neoplastic process in about a quarter of the cases and possibly an epithelial-mesenchymal transition (EMT) in the other cases. Different studies have confirmed that chronic infection with H. pylori induces a chronic inflammation and subsequent damage of the gastric epithelial mucosa, leading to BMDC recruitment. Once recruited, these cells home and differentiate by cell-cell fusion with local gastric epithelial cells, bearing local stem cell failure and participating in tissue regeneration. The context of chronic infection and inflammation leads to an EMT and altered tissue regeneration and differentiation from both local epithelial stem cells and BMDC. EMT induces the emergence of CD44+ cells possessing mesenchymal and stem cell properties, resulting in metaplastic and dysplastic lesions to give rise, after additional epigenetic and mutational events, to the emergence of cancer stem cells (CSCs) and adenocarcinoma.