Neuronal-like cell differentiation of non-adherent bone marrow cell-derived mesenchymal stem cells

Non-adherent bone marrow cell-derived mesenchymal stem cells from C57BL/6J mice were sepa- rated and cultured using the ＂pour-off＂ method. Non-adherent bone marrow cell-derived mesen- chymal stem ceils developed colony-forming unit-fibroblasts, and could be expanded by supple- mentation with epidermal growth factor. Immunocytochemistry showed that the non-adherent bone marrow cell-derived mesenchymal stem cells exposed to basic fibroblast growth factor/epidermal growth factor/nerve growth factor expressed the neuron specific markers, neurofilament-200 and NeuN, in vitro. Non-adherent bone marrow cell-derived mesenchymal stem cells from 13-galactosidase transgenic mice were also transplanted into focal ischemic brain （right corpus striatum） of C57BL/6J mice. At 8 weeks, cells positive for LacZ and 13-galactosidase staining were observed in the ischemic tissues, and cells co-labeled with both 13-galactosidase and NeuN were seen by double immunohistochemical staining. These findings suggest that the non-adherent bone marrow cell-derived mesenchymal stem cells could differentiate into neuronal-like cells in vitro and in vivo.

Contribution of mononuclear bone marrow cells to carbon tetrachloride-induced liver fibrosis in rats

AIM： To study the inhibitory effect of mononuclear bone marrow cell （BNC） transplantation on carbon tetrachloride （CCl4） -induced liver fibrosis in rats. METHODS： Rat liver fibrosis models were induced by CCl4 and alcohol administration. After 8 wk, twenty rats were randomly allocated into treatment group （n = 10） and control group （n = 10）. BMC were infused into the rats in treatment group via the portal vein, while heparinized saline was infused in control group. CCl4 was hypodermically injected into the rats twice a week for 4 wk. At the end of wk 12, all rats were humanely sacrificed. Uver samples were taken and stained with HE or Masson trichrome. The general conditions, liver fibrosis （hydroxyproline and collagen fibre） and liver pathological grades in rats were evaluated. RESULTS： The general conditions of the rats in treatment group improved markedly, but not in control group. Hydroxyproline was 504.6± 128.8 μg/g in treatment group, and 596.0 ± 341.8 μg/g in control group. The percentage of collagen fibre was 3.75% ± 0.98% in treatment group and 5.02% ± 0.44% in control group. There was a significant difference between the two groups （P 〈 0.05）. Liver pathological grade decreased from grade N to grade 11 partially in treatment group （P 〈 0.05） with no obvious improvement in control group （P 〉 0.05）. There was a significant difference between treatment group and control group （P 〈 0.05）.CONCLUSION： Transplantation of BMC can improve liver fibrosis due to chronic liver injury in rats.

Minimally manipulated autologous adherent bone marrow cells (ABMCs):a promising cell therapy of spinal cord injury

Spinal cord injury（SCI）is a devastating ailment that results in drastic life style alterations for the patients and their family members（Mc Donald and Sadowsky,2002）.Damage post injury causes necrosis,edema,hemorrhage and vasospasm.Post injury,secondary damage is caused by ischemia,

Bone marrow cells produce nerve growth factor and promote angiogenesis around transplanted islets

AIM:To clarify the mechanism by which bone marrow cells promote angiogenesis around transplanted islets.METHODS: Streptozotocin induced diabetic BALB/ c mice were transplanted syngeneically under the kidney capsule with the following: (1) 200 islets (islet group: n=12), (2) 1-5×106 bone marrow cells (bone marrow group: n=11), (3) 200 islets and 1-5×106 bone marrow cells (islet + bone marrow group: n= 13), or (4) no cells (sham group:n=5). All mice were evaluated for blood glucose, serum insulin, serum nervegrowth factor (NGF) and glucose tolerance (GTT) up to postoperative day (POD) 14. Histological assessment for insulin, von Willebrand factor (vWF) and NGF was performed at POD 3, 7 and 14.RESULTS: Blood glucose level was lowest and serum insulin was highest in the islet + bone marrow group. Serum NGF increased in islet, bone marrow, and islet + bone marrow groups after transplantation, and there was a significant difference (P=0.0496, ANOVA) between the bone marrow and sham groups. The number of vessels within the graft area was signif icantly increased in both the bone marrow and islet + bone marrow groups at POD 14 as compared to the islet alone group (21.2 ± 3.6 in bone marrow, P=0.01, vs islet group, 22.6 ± 1.9 in islet + bone marrow, P = 0.0003, vs islet group, 5.3 ± 1.6 in islet-alone transplants). NGF was more strongly expressed in bone marrow cells compared with islets. CONCLUSION: Bone marrow cells produce NGF and promote angiogenesis. Islet co-transplantation with bone marrow is associated with improvement of islet graft function.

Diagnostic value of bone marrow cell morphology in visceral leishmaniasis-associated hemophagocytic syndrome:Two case reports

BACKGROUND Visceral leishmaniasis related-hemophagocytic lymphohistiocytosis(VL-HLH)is a hemophagocytic syndrome caused by Leishmania infection.VL-HLH is rare,especially in nonendemic areas where the disease is severe,and mortality rates are high.The key to diagnosing VL-HLH is to find the pathogen;therefore,the Leishmania must be accurately identified for timely clinical treatment.CASE SUMMARY We retrospectively analyzed the clinical data,laboratory examination results,and bone marrow cell morphology of two children with VL-HLH diagnosed via bone marrow cell morphology at Kunming Children’s Hospital of Yunnan,China.Both cases suspected of having malignant tumors at other hospitals and who were unresponsive to treatment were transferred to Kunming Children’s Hospital.They are Han Chinese girls,one was 2 years old and the other one is 9 mo old.They had repeated fevers,pancytopenia,hepatosplenomegaly,hypertriglyceridemia,and hypofibrinogenemia over a long period and met the HLH-2004 criteria.Their HLH genetic test results were negative.Both children underwent chemotherapy as per the HLH-2004 chemotherapy regimen,but it was ineffective and accompanied by serious infections.We found Leishmania amastigotes in their bone marrow via morphological examination of their bone marrow cells,which showed hemophagocytic cells;thus,the children were diagnosed with VL-HLH.After being transferred to a specialty hospital for treatment,the condition was well-controlled.CONCLUSION Morphological examination of bone marrow cells plays an important role in diagnosing VL-HLH.When clinically diagnosing secondary HLH,VL-HLH should be considered in addition to common pathogens,especially in patients for whom HLH-2004 chemotherapy regimens are ineffective.For infants and young children,bone marrow cytology examinations should be performed several times and as early as possible to find the pathogens to reduce potential misdiagnoses.

Therapy with bone marrow cells reduces liver alterations in mice chronically infected by Schistosoma mansoni

AIM： To investigate the potential of bone marrow mononuclear cells （BM-MCs） in the regeneration of hepatic lesions induced by Schistosoma mansoni （S.mansoni） chronic infection. METHODS： Female mice chronically infected with S.rnansoni were treated with BM-MCs obtained from male green fluorescent protein （GFP） transgenic mice by intravenous or intralobular injections. Control mice received injections of saline in similar conditions, Enzyme-linked immunosorbent assay （ELISA） assay for transforming growth factor-beta （TGF-β）, polymerase chain reaction （PCR） for GFP DNA, immunofluorescence and morphometric studies were performed. RESULTS： Transplanted GFP^＋ cells migrated to granuloma areas and reduced the percentage of liver fibrosis. The presence of donor-derived cells was confirmed by Fluorescence in situ hybridization （FISH） analysis for detection of cells bearing Y chromosome and by PCR analysis for detection of GFP DNA. The levels of TGF-β, a cytokine associated with fibrosis deposition, in liver fragments of mice submitted to therapy were reduced. The number of oval cells in liver sections of S.mansoni-infected mice increased 3-4 fold after transplantation. A partial recovery in albumin expression, which is decreased upon infection with S.mansoni, was found in livers of infected mice after cellular therapy. CONCLUSION： In conclusion, transplanted BMCs migrate to and reduce the damage of chronic fibrotic liver lesions caused by S.mansoni.

Induction of Chromosomal Aberrations by Propoxur in Mouse Bone Marrow Cells

Propoxur is a widely used dithiocarbamate insecticide. In this study, the clastogenic effect of propoxur has been evaluated using chromosomal aberration assay in mouse bone marrow cells. Single i. p. administration of propoxur, at 25 mg/kg b.wt., a maximum tolerated dose (MTD) and 12 .5mg/kg b.wt (50% of MTD) have significantly induced different types of aberrations after 24 h of treatment. The aberrations were dose and time dependent and reached a maximum after 24 h of exposure. The sresult suggest a genotoxic potential of propoxur.

The Effect of Sheng Bai Solution on Irradiated Mice Bone Marrow Cell Division Index and DNA Content

Before and after general irradiation with 60Co-γ, mice were orally given Sheng Bai Solution (SBS) for one week. SBS alleviated the irradiation-induced reduction of bone marrow cell chromosome division index. The irradiation-induced decrease of marrow DNA amount, thymic and splenic fractions, and total leukocyte number were restored to some extent. SBS also helped to ameliorate general condition of patients.

An Enzymologic Study on Bone Marrow Cells in Patients with Aplastic Anemia Treated by Supplementing the Kidney and Removing Blood Stasis

The content of cytochrome oxidase (CCO), succinate dehydrogenase (SDH) and neutrophil alkaline phosphatase (NAP) in bone marrow cells in 68 cases of aplastic anemia before and after treatment was determined by computerized graphical analysis and compared with that of normal volunteers (control group). The significantly lowered CCO and SDH levels and the markedly increased NAP content before treatment (P<0.01) became approximately normal after that of supplementing the kidney and removing blood stasis (P >0.05).

Study on Biologic Activity for Membrane of Normal Bone Marrow Cells with Infection of Epidemic Hemorrhagic Fever Virus

Using DPH fluorescence probe, the membrane of normal bone marrow cells with infection of epidemic hemorrhagic fever virus (EHFV) was labeled. The membrane lipid fluidity was obviously decreased from the membrane lipid fluorescence polarization. The membrane lipid fluidity of lympho- cyte, monocyte and neutrophilic granulocyte was dynamically observed. After culturing the cells for 1, 6, 24 and 72 h, it was found that all the membrane lipid fluidity of the infected cells was de- creased obviously with the longer the culturing time, the more obvious it. Compared with the normal control groups, there was a significant difference statistically (P<0. 05-0. 01). It was suggested that the decrease of the membrane lipid fluidity of normal bone marrow cell with infection of EHFV had correlation with the degree of virus invading and cellfunction injury.

Special IgD-λtype multiple myeloma based on bone marrow cell morphology:A case report

We aimed to explore the changes of laboratory indexes of IgD-λtype multiple myeloma with special cell morphology,and to improve the cognition of IgD-λtype MM.To explore the changes of laboratory indexes of IgD-λtype 1 multiple myeloma with special cell morphology,and to improve the cognition of IgD-λtype MM.The morphology of bone marrow cells,immunofixation electrophoresis,serum free light chain(sFLC)and other detection indexes of a patient with IgD-λtype MM treated in Handan Central Hospital in December 2020 were analyzed.The patient bone marrow smears showed 62%of abnormal cells-which were distributed in clusters and resembled lymphoma and metastatic cancer cells.The Flowcytometry indicates that the cell is a plasma cell tumor.Immunoglobulin IgG,IgA and IgM were all lower than the normal range.There is a monoclonal light chainλcomponent in immunofixation electrophoresis.The serum free light chainλwas 2700.00 mg/L,light chain k/λis 0.0023,the high of serum calcium,LDH,β2 microglobulin.IgD-λtype MM is a rare type of MM.The age of onset is young,the invasiveness is strong,the prognosis is poor,the clinical manifestation is complex,and it is easy to be misdiagnosed or missed.The analysis of the clinical symptoms and laboratory characteristics of the disease plays a positive role in the diagnosis,treatment and prognosis of the disease.

The Origin of Neointimal Smooth Muscle Cells in Transplant Arteriosclerosis from Recipient Bone-marrow Cells in Rat Aortic Allograft

In order to investigate the origin of neointimal smooth muscle cells in transplant arterio- sclerosis in rat aortic allograft, sex-mismatched bone marrow transplantation was performed from male Wistar rats to female Wistar rats. Four weeks after transplantation, the aortic transplant model was established by means of micro-surgery in rats. The recipients were divided into 4 groups: female Wistar-female Wistar aortic isografts, female SD-female Wistar aortic allografts, male SD-male Wis- tar aortic allografts, female SD-chimera Wistar aortic allografts. Eight weeks after transplantation, aortic grafts were removed at autopsy and processed for histological evaluation and immunohisto- chemistry. The results indicated that excessive accumulation of α-SMA-positive smooth muscle cells resulted in significant neointima formation and vascular lumen stricture in rat aortic allografts. Neointima assay revealed that the neointimal area and NIA/MA ratio of transplanted artery were sig- nificantly increased in all of aortic allograft groups as compared with those in aortic isograft group (P<0.01). Neointimal smooth muscle cells were harvested from cryostat sections of aortic allograft by microdissection method. The Sry gene-specific PCR was performed, and the result showed that a dis- tinct DNA band of 225 bp emerged in the male-male aortic allograft group and chimera aortic al- lograft group respectively, but not in the female-female aortic allograft group. It was suggested that recipient bone-marrow cells, as the origin of neointimal smooth muscle cells, contributed to the pathological neointimal hyperplasia of aortic allograft and transplant arteriosclerosis.

Bone marrow cell-based regenerative therapy for liver cirrhosis

Bone marrow cells are capable of differentiation into liver cells.Therefore,transplantation of bone marrow cells has considerable potential as a future therapy for regeneration of damaged liver tissue.Autologous bone marrow infusion therapy has been applied to patients with liver cirrhosis,and improvement of liver function parameters has been demonstrated.In this review,we summarize clinical trials of regenerative therapy using bone marrow cells for advanced liver diseases including cirrhosis,as well as topics pertaining to basic in vitro or in vivo approaches in order to outline the essentials of this novel treatment modality.

Bone marrow cell death and proliferation: Controlling mechanisms in normal and leukemic state

Bone marrow cell death and proliferation are regulated by multiple factors including genetic and epigenetic alterations of hematopoietic cells, crosstalk of hematopoietic cells with bone marrow mesenchymal cells through direct cell-cell interaction or cytokine/chemokine production, vascularity of the bone marrow, and interactions of sympathetic nerve system with hematopoiesis. Cell proliferation usually predominates over cell death in neoplastic processes such as leukemia and myeloproliferative neoplasms, while apoptotic processes also have a significant role in the pathogenesis of myelodysplastic syndromes. Recently, hematopoietic stem cells(HSCs) and leukemia stem cells(LSCs) have been identif ied and their characters on self renewal process, differentiation, cell dynamics and drug resistance have been implicated. Although most leukemia cells are initially sensitive to chemo- or radiotherapy, LSCs are resistant and considered to be the basis for disease relapse after initial response. HSCs and LSCs may use similar interactions with bone marrow microenvironment. However, bone marrow microenvironment called niche should inf luence the normal as well as malignant hematopoiesis in different manners. Recent studies have expanded the number of cell types constituting bone marrow niche and made the issue more complex. Since the majority of excellent and contributing studies on bone marrow niches have been performed in animal models, niches in human tissues are beginning to be localized and characterized. In this article, we summarize the relation of hematopoietic cells with niches and hope to point a hint to the novel strategy for treatment of malignant proliferation of hematopoietic cells.

Double potentialities of tumoricide and hematopoiesis of human bone marrow cells activated by IL-2 and IL-3

The biomodulative and hematopoietic potentialities of IL-2 and IL-3 activatedbone marrow(ABM)cells from patients with lung adenocarcinoma were studied in vitro.Human bone marrow(BM)cells could be activated by IL-2 in culture for 7d.TheseIL-2 ABM cells had higher cytolytic activities against cells of H 7402 cell line and freshautologous adenocarcinoma cells and maintained the cytotoxicities longer than IL-2 acti-vated peripheral blood lymphocytes(APBLs),a point of possible importance in adoptiveimmunotherapy for cancer patients.The IL-2 ABM cells also had similar number ofBFU-E and CFU-GM to that had fresh BM cells if 1L-3 was added 48h alter IL-2 inculture.The IL-2 and IL-3 ABM cells might be used to eliminate tumor cells and tosupply reconstitutive elements of BM for autologous bone marrow transplantation.

Comparative study of effects of bone marrow cell vs. Ad_5-HGF administration via non-infarct-related artery injection in myocardial infarction in swine

Objective： To evaluate the effect of transplanting bone marrow-derived mesenchymal stem cells （BM-MSCs） or adenovirus5- hepatocyte growth factor（Ad5-HGF） via non-infarct-related artery injection in swine myocardial infarction models. Methods：BMMSCs were obtained from swine bone marrow and expanded in vitro to a purity of 〉50%. A myocardial infarction（MI） was created by ligating the distal left anterior descending artery in swine. Either BM-MSCs （5 × 10^6/ml） or Ad5-HGF （4 × 10^9 pfu） were transfused via the right coronary artery （non-infarcted artery） four weeks after MI. Gate-controled cardiac perfusion imaging was performed at the end of four and seven weeks after LAD ligation, to evaluate heart function and cardiac perfusion. Morphologic and histologic characteristics of the hearts were also studied. Results： （1）The gate-controlled cardiac perfusion imaging showed that the improvement in LVEF was greater in both treatment groups than in control group at the 4^th weeks. （2）In both treatment groups, capillary density was significantly higher than that of control group（P 〈 0.05）. Conclusion ：BM-MSCs or Ad5-HGF transplantation via non-infarcted artery administration can stimulate angiogenesis and improve heart function, but there was no difference in therapeutic efficacy between BM-MSCs and Ad5-HGF.

Effects of laminin on hard tissue formation by bone marrow cells <i>in vivo</i>and <i>in vitro</i>

The effect of laminin on hard tissue formation using rat bone marrow cells was assessed. Rat bone marrow cells were obtained from femora of 6-week-old male Fischer 344 rats. In this in vivo examination, porous cylindrical hydroxyapatite scaffolds with a hollow center were immersed in 100 mg/ml laminin solution and air-dried. Rat bone marrow cells in 200 ml culture medium at 1 × 106 cells/ml were seeded in the scaffolds. The scaffolds were implanted into the dorsal subcutis of 7-week-old male Fischer 344 rats for 6 weeks. The scaffolds were then removed and examined histologically. For in vitro examinations, 1 × 105 rat bone marrow cells in 2 ml culture medium were then cultured with the addition of dexamethasone and laminin. Rat bone marrow cells were also cultured in laminin-coated culture plates. In vitro examinations showed the effectiveness of laminin for hard tissue formation from the results of biochemical and immunochemical analysis. From the in vivo examination, laminin coating of the scaffolds induced hard tissue in the pores with the cells. It is concluded that laminin is useful for bone formation, as in an in vitro culture study using bone marrow cells, in hydroxyapatite scaffolds in vivo.

Dextran coating on and among fibers of polymer sponge scaffold for osteogenesis by bone marrow cells in vivo

Although hydroxyapatite is commonly used as a scaffold for bone regeneration, sponges may be suitable because of the adaptability to the defect. To use as a scaffold, the fiber of sponge would be coated with any adhesive to storage stem cells in the sponges. Fiber in the structure of commercially available sponges was coated by immersion in dextran solution and air dried. After seeding of rat bone marrow cells (rBMCs), the sponges were implanted subcutis of rats for estimate osteogenesis in vivo. The level of osteocalcin was 25.28 &#177;5.71 ng/scaffold and that of Ca was 129.20 &#177;19.69 μg/scaffold. These values were significantly high- er than those in sponges without dextran coating (p 【0.01). It was thought that rBMCs could be stored on the shelf by dextran deposition in the fiber of the sponge. In vivo examination, dextran induced osteogenesis by rBMCs in many spaces in the inner structure of the sponge.

Effect of L-lysine in culture medium on nodule formation by bone marrow cells

The aim of this study was to estimate the effect of L-lysine on nodule formation by rat bone marrow cells in vitro. In this study, L-lysine was added to medium for mesenchymal stem cell culture to promote proliferation and differentiation of the cells, and then nodule formation was estimated in an in vitro rat bone marrow cell culture. Bone marrow cells from the bone shafts of the femora of Fischer 344 rats were cultured in minimum essential medium with 20 μl of L-lysine solution at 10﹣4, 10﹣5, 10﹣6, 10﹣7 or 10﹣8 M. Dexamethasone was also added to the medium at 10 nM for differentiation of stem cells from bone marrow into osteoblast progenitor cells. The subculture was performed for 2 weeks. The quantity of osteocalcin in rat bone marrow cell culture with dexamethasone was 392 ng/ml. In the medium with dexamethasone and 10﹣8 M L-lysine, the quantity of osteocalcin was 437 ng/ml. Nodules only formed upon addition of 20 μl of L-lysine at 10﹣8 M. It was indicated that 10﹣8 M L-lysine should be the optimal concentration for calcification. For nodule formation by rat bone marrow cells in vitro, the optimum concentration of L-lysine in culture medium might be 20 μl of 10﹣8 M. L-lysine could play an important role in matrix production for bone formation in vitro.

Hybrid Scaffolds Composed of Amino-Acid Coated Sponge and Hydroxyapatite for Hard Tissue Formation by Bone Marrow Cells

A formalin-treated polyvinyl-alcohol (PVF) sponge is convenient as a scaffold because its configuration is easily modified. However, coating the sponge with an adhesive chemical agent is necessary to attach bone marrow cells (BMCs) to the sponge structure. Moreover, it was considered that a hybrid scaffold composed of a sponge and enveloped cylindrical porous hydroxyapatite (HA) would be convenient. In this study, the effect of leucine (Leu) coating on a PVF sponge was examined for osteogenesis on an HA/PVF hybrid scaffold by rat BMCs (rBMCs). In an in vivo assessment, the sponge immersed in Leu solution (10 mg/ml) was inserted into the hollow center of cylindrical HA. The sponge received 1.5 × 106 rBMCs obtained from male Fischer 344 rats. The hybrid scaffolds were then implanted subcutaneously of syngeneic rats for 6 weeks. In vitro assessment of Leu to hard tissue formation with coating on the well or addition in rBMC culture medium was also performed in a 6-well plate for 2 weeks. In vivo examinations showed the excellent effect of Leu coating on PVF sponge. Leu-coated PVF sponge in the scaffolds showed marked new bone formation in the pores by histological examination. Leu-coated PVF sponge showed a high quantity of osteocalcine (OC). HA might prevent the release of rBMCs from PVF as a barrier. In in vitro examinations, the quantity of OC in rBMC culture with and without the addition of Leu in culture medium showed no significant difference. However, addition of Leu showed significant ALP activity level in culture medium. Leu coating in culture plate wells showed no influence on the quantity of OC. It was concluded from the results that Leu might prevent the emigration of rBMCs to the outside of the scaffold and promote the differentiation of cells to osteoblasts in the scaffold.