Rat Bone Marrow Cells Research Articles

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 ± 5.71 ng/scaffold and that of Ca was 129.20 ± 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.

Functional Impact of Targeted Closed-Chest Transplantation of Bone Marrow Cells in Rats with Acute Myocardial Ischemia/Reperfusion Injury

Intramyocardial transplantation of bone marrow-derived stem cells is a potential therapeutic option after myocardial infarction (MI). Intramyocardial administration is invasive but allows efficient and targeted stem cell delivery. Aims of this study were validation of minimal-invasive, echo-guided closed-chest cell transplantation (CTx) of mononuclear (MNC) or mesenchymal stem cells (MSC) and quantification of systolic left ventricular function and assessment of contractile reserve with high-resolution reconstructive 3D-echocardiography (r3D-echo) 3 weeks after CTx. Female Fischer344 rats received syngeneic male MNC, MSC, or medium after myocardial ischemia and reperfusion via echo-guided percutaneous injection (open-chest for control). Left ventricular systolic function was measured and dysfunctional myocardium was quantified with r3D-echo. For investigation of contractile reserve and myocardial viability r3D-echo was additionally conducted during low-dose dobutamine 3 weeks after CTx. Cell persistence after echo-guided CTx was quantified via real-time PCR; scar size was measured histologically. Echo-guided percutaneous CTx was feasible in all animals (n = 30) without periprocedural complications. After 3 weeks, 1.4 +/- 1.1% of transplanted MNC and 1.9 +/- 1.2% of MSC were detected. These numbers were comparable to those after open-chest intramyocardial injection of MNC (0.8 +/- 1.1%; n = 8, p = 0.3). In r3D-echo no functional benefit was associated with CTx after MI and reperfusion. All groups (MNC, MSC, and controls) revealed a significant decrease of dysfunctional myocardium and similar contractile reserve during inotropic stimulation.In conclusion, percutaneous echo-guided closed-chest CTx promises to be an effective and safe approach for CTx in small-animal research. However, intramyocardial CTx of MNC or MSC had no influence on systolic function and contractile reserve after reperfused MI.

Anticlastogenic effect of caffeic acid phenethyl ester on cisplatin-induced chromosome aberrations in rat bone marrow cells

Caffeic acid phenethyl ester (CAPE) is an antioxidant that can scavenge free radicals and protect cellular macromolecules, including DNA and proteins, from oxidative damage induced by various agents. The protective effect of CAPE on cisplatin-induced chromosome aberrations has been determined in rat bone marrow cells. The animals were pretreated with a single dose of CAPE (10 micromol/kg body weight [b.w.]) injected intraperitoneally (i.p.) 24 hours before the administration of cisplatin and then sacrificed 24 hours after the cisplatin administration. Cisplatin was administered to rats either alone (5 mg/kg b.w., i.p.) or after CAPE treatment. CAPE has led to a statistically significant decrease in the total number of chromosomal aberrations and abnormal metaphases induced by cisplatin when compared with only cisplatin given groups. We have concluded that CAPE could prevent cisplatin-induced chromosome aberrations by establishing a potent free radical scavenger effect.

Evaluation of the genotoxic potential of Austroplenckia populnea (Reiss) Lundell chloroform fraction from barkwood extract in rodent cells in vivo

The genotoxic effect of the Austroplenckia populnea chloroform fraction from barkwood extract was tested in vivo on peripheral blood cells of Swiss mice with the comet assay (SCGE), and the clastogenic effect was investigated on peripheral blood cells of Swiss mice and bone marrow cells of Wistar rats, with the micronucleus and chromosome aberrations tests. The animals were treated by gavage with 3 concentrations of the extract: 300, 600 and 900 mg.kg-1. Peripheral blood cells of Swiss mice were collected 4 and 24 hours after the treatment to the SCGE assay and 48 and 72 hours to the micronucleus test. Bone marrow cells of Wistar rats were collected 24 hours after the treatment to the micronucleus and chromosome aberration tests. The results showed that the A. populnea chloroform fraction induced an increase in the average number of DNA damage in peripheral blood cells at the three concentrations tested, but this increase was not statistically significant. In the micronucleus and chromosome aberrations test, no significant increase was observed in the mean number of micronucleated polychromatic erythrocytes (MNPCE) of Swiss mice or MNPCE or chromosome aberrations for the rat bone marrow cells, for any of the tested doses. Our findings enable us to conclude that by the comet assay, A. populnea chloroform fraction from barkwood extract showed no genotoxic effects, and by the micronucleus and chromosome aberration tests, the extract fraction showed no clastogenic/aneugenic effects on the rodent cells tested.

Tranilast suppresses prostate cancer growth and osteoclast differentiation in vivo and in vitro

In bone metastatic sites, prostate cancer cells proliferate on interacting with osteoclasts. Tranilast, which is used for an antiallergic drug, has been shown to inhibit growth of several cancers and stromal cells. The present study was conducted to assess suppressive effects of Tranilast on prostate cancer growth and osteoclast differentiation in vivo and in vitro. In vivo, rat prostate cancer tissue was transplanted onto cranial bones of F344 rats and Tranilast was given for 9 days at doses of 0, 200, or 400 mg/kg/day. In vitro, human prostate cancer cell lines, LNCaP, PC3, and DU145, the rat prostate cancer cell line, PLS-10, and rat bone marrow cells were similarly treated with the agent. In vivo, tumor volumes were significantly decreased in the high dose group. While cell proliferation did not appear to be affected, apoptosis was induced and tumor necrosis was apparent. Cranial bone defects were decreased in the high dose group. In vitro, cell proliferation rates of all four cell lines were reduced by Tranilast and increased apoptosis was observed in LNCaP and PLS-10. In addition, Tranilast significantly reduced osteoclast differentiation of rat bone marrow cells. Western blot analysis of PLS-10 and LNCaP revealed that phospho-GSK3beta was up-regulated and phospho-Akt was down-regulated. Tranilast here suppressed rat prostate cancer growth and osteoclast differentiation. Growth of human prostate cancer cells was also inhibited. Thus, this agent deserves consideration as a candidate for conventional therapy of bone metastatic prostate cancer.

Assessment of the mutagenic effect of 1,1-dimethyl hydrazine

The mutagenic effect of the rocket fuel 1,1-dimethyl hydrazine has been studied experimentally and compared to the well-recognized mutagene N-nitroso dimethylamine. The manifestation of the effect for both compounds was disclosed through a significant increase in the chromosome aberration frequency in the bone marrow cells of intoxicated rats. The levels of chromosome aberrations induced by 1,1-dimetyl hydrazine were studied following both single (1 h) and repeated doses (daily for 10 consecutive days) by inhalation (205–1028 mg/m 3) and gavage (5.4–26.8 mg/kg) administration, respectively. For comparison N-nitroso dimethylamine were administered by inhalation (2 h/daily for 10 consecutive days) and by gavage in concentrations of 2.4–48 mg/m 3 and 1–30 mg/kg, respectively. A clear dependence of concentration as well of time was disclosed. The BenchMark Dose approach was employed to derive guideline doses for the two compounds, the implications towards human health being discussed.

Raloxifene: its ossification-promoting effect on female mesenchymal stem cells

BackgroundRaloxifene acts like estrogen in preventing bone loss in postmenopausal women, but it selectively activates biological responses in bone tissue. It has a direct effect on osteoblasts’ differentiation and bone formation in bone marrow culture. However, the point at which raloxifene has an effect on bone marrow-derived mesenchymal stem cells (MSCs), regardless of sex difference, is not known. The purpose of this study was to examine the osteogenic effect of raloxifene on MSCs derived from female and male rats and to assess the sex difference of raloxifene with or without osteogenic supplements (OSs) in the regulation of bone formation. MethodsFemale and male rat bone marrow cells were cultured with or without OSs. In each experimental group, 106 M or 10-8 M raloxifene was added. As a control, cells were cultured without raloxifene. Histologically, mineralization was assessed by alizarin red S staining. Biochemically, alkaline phosphatase (ALP) activity, calcium content, and osteocalcin content were assessed. ResultsOn histological analysis, mineralized nodules were seen on alizarin red S staining in the groups treated with OS. On the biochemical analysis, OS increased ALP activity, calcium content, and osteocalcin content. Among female groups with OSs, 10-6 M raloxifene significantly increased ALP activity, calcium content, and osteocalcin content compared with the controls. Among male groups, raloxifene had negligible effects. Conclusions10-6 M Raloxifene had no ossification-inducing effect on female MSCs, but it had an ossification-promoting effect; it had no osteogenic effect on male MSCs. Therefore, raloxifene has a sex difference with regard to its osteogenic effect on MSCs. Moreover, combined treatment with raloxifene plus OS has an effect on female MSCs. These results provide a useful insight into the possible influence of raloxifene after MSC transplantation in clinical practice.

Cell sheet transplantation of cultured mesenchymal stem cells enhances bone formation in a rat nonunion model

Orthopedic surgeons have long been troubled by cases involving nonunion of fractured bones. This study aimed to enhance bone union by cell sheet transplantation of mesenchymal stem cells. A nonunion model was made in rat femur, and rat bone marrow cells were cultured in medium containing dexamethasone and ascorbic acid phosphate to create a cell sheet that could be scraped off as a single sheet. Cell sheets were transplanted onto fractured femurs without a scaffold in the model. X-ray and histological analysis were performed at 2, 4 and 8 weeks. Ultrasonography and biomechanical analysis were performed at 8 weeks. X-ray photographs and histological sections showed callus formation around the fracture site in the cell sheet-transplanted group (sheet group). Bone union was obtained in the sheet group at 8 weeks. By contrast, the control group (without sheet transplantation) showed nonunion of the femur. The results of pullout evaluation in the vertical direction of the femur in the sheet group were significantly better than that of the control group. Analysis of the origin of de novo formed bone using the Sry gene, which was used as a marker for donor cells, showed that transplanted cells without scaffolds could survive and differentiate into osteogenic lineage cells in vivo. These results showed that the femoral fracture in our model was completely cured by the transplantation of a cell sheet created by tissue engineering techniques. Thus, we think that cell sheet transplantation can contribute to hard tissue reconstruction in cases involving nonunion, bone defects and osteonecrosis.

Ozone inhalation can induce chromosomal abnormalities in bone marrow cells of Wistar rats

Due to the wide range of applications for ozone and its increasing use for medical and industrial purposes, studying its effects has become a very important line of research. The ozone has been suspected to be a carsinogen. Because of the increasing use of ozone, the human could be more and more exposed to this gas. In this study the effects of ozone inhalation on chromosomes and its clastogenic consequences have been investigated using in vivo micronucleus assay in bone marrow cells of treated rats. Animals were treated for 6 hours a day at 3 ppm of ozone during 10 consecutive days. The micronucleus assay was performed immediately and 11 days after the last exposure. The frequency of micronucleated polychromatic erythrocyte of bone marrow (MNPCE) increased in both groups compared to the control. Such increase confirmed the clastogenic effects of ozone. The elevated frequency of MNPCE did not decrease after 11 days of the last ozone exposure. Results indicate that ozone inhalation could induce persistent chromosomal damages even to bone marrow cells which were not in direct contact to it. Also, once more, the results confirmed the usefulness of the micronucleus assay in toxicological studies.

Osteogenic and adipogenic differentiation of rat bone marrow cells on non-mulberry and mulberry silk gland fibroin 3D scaffolds

This study investigates the potential of 3D silk scaffolds fabricated using tropical tasar non-mulberry, Antheraea mylitta and mulberry, Bombyx mori silk gland fibroin proteins as substrate for osteogenic and adipogenic differentiation of rat bone marrow cells (BMCs). The scaffolds are mechanically robust and show homogenous pore distribution with high porosity and interconnected pore walls. Low immunogenicity of fabricated silk scaffolds as estimated through TNF α release indicates its potential as future biopolymeric graft material. Rat bone marrow cells cultured on scaffolds for 28 days under static conditions in osteogenic and adipogenic media respectively led to induction of differentiation. Proliferation and spreading of fibroblasts and bone marrow cells on silk scaffolds were observed to be dependent on scaffold porosity as revealed through confocal microscopic observations. Histological analysis shows osteogenic differentiation within silk scaffolds resulting in extensive mineralization in the form of deposited nodules as observed through intense Alizarin Red S staining. Similarly, adipogenesis was marked by the presence of lipid droplets within scaffolds on staining with Oil Red O. Real-time PCR studies reveal higher transcript levels for osteopontin ( Spp1), osteocalcin ( Bglap2) and osteonectin ( Sparc) genes under osteogenic conditions. Similarly, upregulated adipogenic gene expression was observed within A. mylitta and B. mori scaffolds under adipogenic conditions for Peroxisome proliferator activated receptor gamma ( PPARγ2), lipoprotein lipase ( LPL) and adipocyte binding protein ( aP2) genes. The results suggest suitability of silk fibroin protein 3D scaffolds as natural biopolymer for potential bone and adipose tissue engineering applications.

Insulin receptor substrate 2 plays important roles in 17β-estradiol-induced bone formation

Discovering the mechanisms of the estrogen effects on the osteoblasts is very important for the development of new agents which have the clear-cut beneficial effects of estrogen while free of adverse effect. The aim of this study was to investigate the differential gene expression of 17beta-estradiol (E2)-treated osteoblast-like cells, and the effect of E2 on the insulin receptor substrate 2 (IRS- 2) expression in human cultured osteoblast-like cells and the osteoblasts of ovariectomized (OVX) rats. The differential gene expression of E2-treated osteoblast- like cells was analyzed by cytokine expression array and validated by RT-PCR and Western blot analysis. The protein expression and phosphorylation of one of the differentially expressed gene, IRS-2, treated at different times with E2 were analyzed. The Sprague-Dawley rats were ovariectomized and then treated with E2, the IRS-2 expression was analyzed by immunohistochemistry analysis. E2 upregulated the mRNA expression of IRS-2, bone morphogenetic protein 9, and connective tissue growth factor expression, down-regulated the mRNA expression of matrix metalloproteinase 15 and some tumor suppressor genes. Peak expression of IRS-2 was observed at 12-24 h of treatment by 10-8M E2. E2 can also increase the phosphorylation of IRS-2. The IRS-2 expression was down-regulated in the osteoblasts and bone marrow cells of the OVX rats, which had lower bone mineral density (BMD) than the normal rats. However, both BMD and IRS-2 expression can be rescued by 10-8M E2 in the OVX rats. IRS-2 in osteoblast is up-regulated by E2 and plays important roles in the estrogen- induced bone formation.

Role of Nbr1 in bone remodeling

The growth and differentiation of bone cells is controlled by various factors which can be modulated by heparan sulfates. Here, we investigated the effects of a low-molecular-weight “heparin-like”, an oversulfated exopolysaccharide (OS-EPS) produced by a mesophilic strain found in deep-sea hydrothermal vents, on bone cells in regard to their differentiation, proliferation, and function in vitro. We compared this effect with that of a non-oversulfated exopolysaccharide (EPS). Cell viability was assessed by trypan blue exclusion, and alive and dead cells were manually counted. Besides, by time-lapse experiments, the cell divisions were manually scored. Mesenchymal stem cells (MSCs) isolated and propagated previously from the rat bone marrow were tested for their capabilities to differentiate into osteoblasts in vitro. We showed that exogenous application of OS-EPS to cultures of primary MSCs during in vitro differentiation inhibited MSC mineralization. In addition, OS-EPS strongly reduces the proliferation of differentiating osteoblasts MSCs as we observed for the rat bone marrow cells. This proliferation inhibition was not due to an induction of apoptosis. These results suggest that OS-EPS reduces the differentiated cells proliferation, leading to strong inhibition of mineralized nodule formation. In contrast, OS-EPS enhanced no differenciated MSCs proliferation. EPS had no effect in any of the experimental conditions, underlying the importance of sulfatation in the proliferation and differentiation effects. Bone remodelling is regulated by osteogenic cells which act individually through cellular and molecular interaction. Osteoclasts are bone-resorptive cells that differentiate from hematopoietic precursors upon receptor activator of nuclear factor kappaB ligand (RANKL) activation. To investigate the OS-EPS effect in osteoclastic differentiation, we used several models: murine RAW264.7 cells and human CD14+ monocytes. In the two models used, the addition of OS-EPS inhibited RANKL-induced osteoclastogenesis. In contrast to EPS or other glycosaminoglycan, surface plasmon resonance experiments revealed that RANKL and OS-EPS are able to form a complex. In addition, a strong decrease of the osteoclastic precursor cells adhesion was observed. These results suggest that OS-EPS is a strongly inhibitor of osteoclastogenesis. It also underlines the importance of OS-EPS as a co-receptor for the signaling of RANKL. Conflict of interest: None declared.

Rat marrow‐derived mesenchymal stem cells developed in a medium supplemented with the autologous versus bovine serum

The controversial effect of autologous serum (AS) on human mesenchymal stem cells (MSC) was studied in rat MSC culture. Rat bone marrow cells were plated in a medium containing either FBS (fetal bovine serum) or AS were cultured to passage 3, during which the population doubling number (PDN) of both cultures were measured and compared statistically. The number of viable cells, the cell colonogic activity, and cell growth rate were also compared. In addition, mineralization in the osteogenic cultures from each system was measured. Our data indicated that AS enriched medium provided a microenvironment in which growth rate as well as bone differentiation of the isolated MSCs were significantly higher than in FBS enriched medium.

Effects of berberine on differentiation and bone resorption of osteoclasts derived from rat bone marrow cells

To observe the effects of berberine on osteoclastic differentiation and bone resorption action in vitro, and to investigate the cellular mechanism of its inhibitory effects on bone resorption. The multinucleated osteoclasts (MNCs) were derived by 1,25-dihydroxyvitamin D3 and dexamethasone from bone marrow cells in the coculture system with primary osteoblastic cells. The tartrate-resistant acid phosphatase (TRAP) staining and image analysis of bone resorption pit on dental slices were used to identify osteoclast. The activity of TRAP was measured by p-nitrophenyl sodium phosphate assay. The bone resorption pit area on the bone slices formed by osteoclasts was measured by computer image processing. At the concentrations of 0.1, 1 and 10 micromol/L, berberine dose-dependently suppressed the formation of TRAP-positive multinucleated cells, the TRAP activity and the osteoclastic bone resorption. The strongest inhibitory effect was exhibited at the concentration of 10 micromol/L, with the inhibiting rate of 60.45%, 42.12% and 72.69% respectively. Berberine can decrease bone loss through inhibition of osteoclast formation, differentiation and bone resorption.

Antigenotoxic effect of genistein against 7,12-dimethylbenz[a]anthracene induced genotoxicity in bone marrow cells of female wistar rats

Carcinogen induced mutation in somatic cells leads to genetic instability, which is considered as an important facet of carcinogenesis. Agents that inhibit DNAadduct formation, stimulate DNArepair mechanisms, and possess antioxidant functions are considered as antigenotoxic agents. Genistein, the major isoflavone of soy products, protects animals against experimentally induced mammary and prostate cancers. 7,12-dimethylbenz[a]anthracene (DMBA), a potent site-specific carcinogen, induce mutations in DNA through its active metabolite, dihydrodiol epoxide, is a crucial step in cancer initiation. The antigenotoxic effect of genistein against DMBA-induced genotoxicity has been investigated in the present study by analyzing the frequency of micronucleated polychromatic erythrocytes (MnPCEs) and chromosomal aberrations as cytogenetic end-points. The status of lipid peroxidation, antioxidants and detoxication agents were used as biochemical end-points to assess the antigenotoxic effect of genistein. Elevated MnPCEs frequency, marked chromosomal aberrations and enhanced status of lipid peroxidation, antioxidants and detoxication agents were observed in DMBAtreated animals. Oral pretreatment of genistein (20mg/kg b.w.) for 5 days to DMBAtreated animals significantly reduced the frequency of micronucleus formation and chromosomal abnormalities as well as reversed the status of biochemical variables. Our results suggest that genistein has potent antigenotoxic effect against DMBA-induced genotoxicity.

Platelet‐rich plasma suppresses osteoclastogenesis by promoting the secretion of osteoprotegerin

Platelet-rich plasma is characterized by containing fundamental protein growth factors. Although many in vitro studies have documented the capability of platelet-rich plasma to induce the growth of osteoblasts or osteoblast-like cells, the effect of platelet-rich plasma on osteoclastogenesis has not yet been studied. The aim of the present study was to evaluate the effects of platelet-rich plasma and platelet-poor plasma on osteoclastogenesis with rat bone marrow cell culture. Platelet-rich plasma and platelet-poor plasma were produced from the whole blood of rat. For cell culture, rat bone marrow cells were isolated from rat tibiae and then treated with 1,25alpha dihydroxy vitamin D(3) and with different concentrations of platelet-rich plasma or platelet-poor plasma. After 4 d of culture, rat bone marrow cells were stained with tartrate-resistant acid phosphatase (TRAP), and TRAP-positive cells that had more than three nuclei (TRAP-positive multinucleated cells) were counted as osteoclast-like cells. Osteoprotegerin, known as an osteoclastogenesis-related factor, cells was quantified using an enzyme-linked immunosorbent assay (ELISA). Although platelet-poor plasma had no effect on the formation of TRAP-positive multinucleated cells, platelet-rich plasma decreased the number of TRAP-positive multinucleated cells in a dose-dependent manner. The amount of osteoprotegerin produced from rat bone marrow cells and from MC3T3-E1 cells was enhanced in platelet-rich plasma-treated groups. Under our experimental conditions, platelet-rich plasma decreased the formation of TRAP-positive multinucleated cells and increased the secretion of osteoprotegerin. This study suggests that platelet-rich plasma suppresses osteoclastogenesis, therefore inhibiting bone resorption. In addition we also demonstrated that platelet-rich plasma increased the secretion of osteoprotegerin, an inhibitor for osteoclast formation, thus suggesting that the enhancement of osteoprotegerin secretion induces this inhibitory effect.

Near-optimal Conditions for the In Vitro Culture of Hemopoietic Progenitor Cells in Bone Marrow from the Rat

In vitro techniques for the culture of hemopoietic stem cells and committed hemopoietic progenitor cells in rat bone marrow have not been adequately described in the literature. In the present investigations, and using commercially available hemopoietic cytokines and growth factors, the conditions required to perform long-term bone marrow culture (LTBMC) using rat femoral bone marrow were studied, in conjunction with the colony-forming unit cell assay (CFU-C), to quantify the number of progenitor cells. CFU-C production by LTBMCs, set up using Iscove's modified Dulbecco's medium supplemented with fetal calf serum and horse serum, ceased after week 2 of culture. However, the duration of CFU-C production was significantly increased by supplementing LTBMCs with the cytokine recombinant mouse stem cell factor or recombinant rat stem cell factor.

Effect of Every-Other-Day Fasting on Spontaneous Chromosomal Damage in Rat's Bone-Marrow Cells

Dietary restriction in experimental rodents, either by calorie restriction (CR) or by every-other-day fasting (EODF), was shown to protect against cancer and increase lifespan. One of the suggested hypotheses to explain the beneficial effects of dietary restriction is that the diet stabilizes the integrity of the genetic information. The effects of EODF on the spontaneous frequency of sister chromatid exchanges (SCE) and chromosomal aberrations (CA) were examined in bone-marrow cells of 3-mo-old Wistar male rats. After 12 wk of EODF diet, significant reduction in the frequency of SCE and total number of CA was observed. Data indicate a protective effect of EODF diet against spontaneous mutations in rats.

Effects of Bone Marrow and Hepatocyte Transplantation on Liver Injury

The therapeutic effects of bone marrow and hepatocyte transplantation were investigated regarding the treatment of retrorsine-partial hepatectomy-induced liver injury. Analbuminemic F344alb rats were given two doses of retrorsine 2 wk apart, followed 4 wk later by transplantation with F344 rat bone marrow cells or hepatocytes immediately after a two-thirds hepatectomy. The survival rate, liver regeneration rate, liver functions, albumin-positive hepatocytes, and normal albumin gene sequences in the liver and serum albumin levels were investigated in the recipients. Although 65% retrorsine/partial hepatectomy-treated F344alb died between 1 and 11 d after the partial hepatectomy, only 27.5% of the animals died following bone marrow transplantation, and 50% with hepatocyte transplantation. Both bone marrow and hepatocyte transplantation ameliorated acute liver injury after a partial hepatectomy. Bone marrow transplantation yielded a very small increase in the number of albumin-positive hepatocytes in the liver, while hepatocyte transplantation resulted in massive replacement of the liver tissues by the donor hepatocytes associated with an elevation of serum albumin after an extended time. Both bone marrow and hepatocyte transplantation could prevent acute hepatic injury, conceivably due to a paracrine mechanism.

Passage of bone-marrow-derived liver stem cells in a proliferating culture system

To explore the feasibility of passage of bone-marrow-derived liver stem cells (BDLSCs) in culture systems that contain cholestatic serum. Whole bone marrow cells of rats were purified with conditioning selection media that contained 50 mL/L cholestatic serum. The selected BDLSCs were grown in a proliferating culture system and a differentiating culture system. The culture systems contained factors that stimulated the proliferation and differentiation of BDLSCs. Each passage of the proliferated stem cells was subjected to flow cytometry to detect stem cell markers. The morphology and phenotypic markers of BDLSCs were characterized using immunohistochemistry, reverse transcription polymerase chain reaction (RT-PCR) and electron microscopy. The metabolic functions of differentiated cells were also determined by glycogen staining and urea assay. The conditioning selection medium isolated BDLSCs directly from cultured bone marrow cells. The selected BDLSCs could be proliferated for six passages and maintained stable markers in our proliferating system. When the culture system was changed to a differentiating system, hepatocyte-like colony-forming units (H-CFUs) were formed. H-CFUs expressed markers of embryonic hepatocytes (alpha-fetoprotein, albumin and cytokeratin 8/18), biliary cells (cytokeratin 19), hepatocyte functional proteins (transthyretin and cytochrome P450-2b1), and hepatocyte nuclear factors 1alpha and -3beta). They also had glycogen storage and urea synthesis functions, two of the critical features of hepatocytes. BDLSCs can be selected directly from bone marrow cells, and pure BDLSCs can be proliferated for six passages. The differentiated cells have hepatocyte-like phenotypes and functions. BDLSCs represent a new method to provide a readily available alternate source of cells for clinical hepatocyte therapy.