Biochemical and molecular characterization of hepatocyte-like cells derived from human bone marrow mesenchymal stem cells on a novel three-dimensional biocompatible nanofibrous scaffold

Background:  There is significant interest in using nanofibers in tissue engineering from stem cells. The transdifferentiation of mesenchymal stem cells into the hepatic lineage in a nanofibrous structure has not been reported. In this study, a three dimensional nanofibrous scaffold is introduced for differentiation of human bone marrow derived mesenchymal stem cells (hBMSCs) into hepatocytes.
Methods:  A scaffold composed of Poly (ε-caprolactone), collagen and polyethersulfone was fabricated by the electrospinning technique. After characterization of isolated hBMSCs, the performance of the cells on the scaffold was evaluated by Scanning Electron Microscopy (SEM) and MTT assay. Cytological, molecular and biochemical markers were measured to confirm differentiation potential of hBMSCs into hepatocytes.
Results:  The isolated cells possessed the basic properties of mesenchymal stem cells (MSCs). Based on scanning electron microscope (SEM) analysis and MTT assay, it was shown that the cells adhere, penetrate and proliferate on the nanofibers. Cultured cells on the nanofibers differentiated into hepatocyte-like cells and expressed hepatocyte specific markers such as albumin, α-fetoprotein, cytokeratin-18, cytokeratin-19 and cytochrome P450 3A4 at mRNA levels. Appearance of a considerable number of albumin-positive cells cultivated on the scaffold (47 ± 4%) as compared to the two-dimensional culture system (28 ± 6%) indicates the supporting role of the scaffold. The efficiency of the cells to produce albumin, urea, transferrin, serum glutamic pyruvic transaminase and serum oxaloacetate aminotransferase in hepatocytes on the scaffold further attest to the functionality of the cells.
Conclusion:  The data presented in this study show that the engineered nanofibrous scaffold is a conductive matrix which supports and enhances MSC development into functional hepatocyte-like cells.     

Efifcient generation of functional hepatocyte-like cells from mouse liver progenitor cells via indirect co-culture with immortalized human hepatic stellate cells

BACKGROUND: Differentiation of liver progenitor cells (LPCs) to functional hepatocytes holds great potential to de-velop new strategies for hepatocyte transplantation and the screening of drug-induced cytotoxicity. However, reports on the efifcient and convenient hepatic differentiation of LPCs to hepatocytes are few. The present study aims to investigate the possibility of generating functional hepatocytes from LPCs in an indirect co-culture system.
METHODS: Mouse LPCs were co-cultured in Transwell plates with an immortalized human hepatic stellate cell line (HSC-Li) we previously established. The morphology, expression of hepatic markers, and functions of mouse LPC-derived cells were monitored and compared with those of conventionally cultured LPCs.
RESULTS: Co-culturing with HSC-Li cells induced differentia-tion of mouse LPCs into functional hepatocyte-like cells. The differentiated cells were morphologically transformed into hepatocyte-like cells 3 days after co-culture initiation. In addi-tion, the differentiated cells expressed liver-speciifc genes and possessed hepatic functions, including glycogen storage, low-density lipoprotein uptake, albumin secretion, urea synthesis, and cytochrome P450 1A2 enzymatic activity.
CONCLUSIONS: Our method, which employs indirect co-culture with HSC-Li cells, can efifciently induce the differentiation of LPCs into functional hepatocytes. This ifnding suggests that this co-culture system can be a useful method for the efifcient generation of functional hepatocytes from LPCs.     

A randomized study of buffy coat platelets in platelet additive solution stored 1–5 versus 6–7 days prior to prophylactic transfusion of allogeneic haematopoietic progenitor cell transplant recipients

Background and Objective  Storage of platelets > 5 days provides improved availability, logistical management and decreased outdating. Promising results on in vitro parameters and on in vivo post-transfusion recovery and survival of autologous platelets in healthy volunteers have earlier been shown. To provide additional verification, randomized patient transfusion studies are needed.
Materials and Methods  Sixty allogeneic haematopoietic progenitor cell transplant recipients were randomized to receive buffy-coat (BC) platelets stored in platelet additive solution (PAS) for 1–5 days the first time a prophylactic transfusion was needed after transplantation, followed the second time by platelets stored for 6–7 days or vice versa. The corrected count increment (CCI) for 1 and 24 h were calculated.
Results  CCI 1 h and CCI 24 h were higher for platelets stored 1–5 days as compared to 6–7 days, 10·4 ± 5·1 vs. 7·4 ± 3·8 ( P <  0·001) and 5·4 ± 4·1 vs. 2·6 ± 2·6 ( P <  0·001), respectively. Time to next platelet transfusion was significantly longer after a transfusion of platelets stored for 1–5 days as compared to platelets stored for 6–7 days: 2·2 ± 1·1 vs. 1·6 ± 0·8 days, respectively ( P <  0·005). No differences in bleeding events and no transfusion reaction were recorded.
Conclusion  The advantage of an extension of platelet storage time beyond day 5 should be balanced against the increased need for platelet transfusions that may occur and the conceivable risk of transfusion failure.     

Immediate intraportal transplantation of human bone marrow mesenchymal stem cells prevents death from fulminant hepatic failure in pigs

The effectiveness of human bone marrow mesenchymal stem cell (hBMSC) transplantation to treat acute and chronic liver injury has been demonstrated in animal models and in a few nonrandomized clinical trials. However, no studies have investigated hBMSC transplantation in the treatment of fulminant hepatic failure (FHF), especially in large animal (pig) models. The aim of this study was to demonstrate the safety, effectiveness, and underlying mechanism of hBMSC transplantation for treating FHF in pigs through the intraportal route. Human BMSCs (3 × 10(7) ) were transplanted into pigs with FHF via the intraportal route or peripheral vein immediately after D-galactosamine injection, and a sham group underwent intraportal transplantation (IPT) without cells (IPT, peripheral vein transplantation [PVT], and control groups, respectively, n = 15 per group). All of the animals in the PVT and control groups died of FHF within 96 hours. In contrast, 13 of 15 animals in the IPT group achieved long-term survival (>6 months). Immunohistochemistry demonstrated that transplanted hBMSC-derived hepatocytes in surviving animals were widely distributed in the hepatic lobules and the liver parenchyma from weeks 2 to 10. Thirty percent of the hepatocytes were hBMSC-derived. However, the number of transplanted cells decreased significantly at week 15. Only a few single cells were scattered in the regenerated liver lobules at week 20, and the liver tissues exhibited a nearly normal structure. Conclusion: Immediate IPT of hBMSCs is a safe and effective treatment for FHF. The transplanted hBMSCs may quickly participate in liver regeneration via proliferation and transdifferentiation into hepatocytes during the initial stage of FHF. This method can possibly be used in future clinical therapy. (HEPATOLOGY 2012;56:1044-1052).     

损伤肝脏条件培养液体外诱导小鼠骨髓间充质干细胞分化为肝细胞

目的探索损伤肝脏条件培养液体外诱导小鼠骨髓间充质干细胞（MSCs）分化为肝细胞的可行性及方法。方法培养注射CCl4所致的损伤小鼠肝脏组织块,收集条件培养液,进行MSCs诱导分化。通过形态学观察、RT-PCR、免疫荧光反应和过碘酸Schiff反应鉴定分化细胞。结果诱导组细胞呈肝细胞样变化。RT-PCR检测显示：AFP基因第5天开始表达,第10天表达增强,此后表达减弱；细胞角蛋白18和Alb基因第10天开始表达,持续到第20天；第20天检测到酪氨酸氨基转移酶基因表达。免疫荧光反应显示诱导20 d的细胞AFP、细胞角蛋白18、Alb表达阳性,诱导20 d细胞过碘酸Schiff反应呈阳性。结论MSCs在损伤肝脏条件培养液诱导下可分化为肝细胞。     

Platelet vascular endothelial growth factor is a useful predictor for prognosis in Kawasaki syndrome

Kawasaki syndrome (KS) is an acute febrile vasculitis of childhood. Coronary artery abnormalities (CAA) are a significant problem in KS patients. High dose intravenous immunoglobulin (IVIG) is effective for reducing the occurrence of CAA. Clinical and histopathological findings suggest that vascular endothelial growth factor (VEGF) is involved in CAA. In circulating blood, newly activated platelets are the major source of VEGF, which is released in large amounts in vascular inflammation. The present study analysed 80 KS patients (69 IVIG responders and 11 IVIG non-responders) and evaluated the role of platelet VEGF in KS vasculitis. Serum VEGF and platelet VEGF levels were significantly higher in KS patients than controls ( P  <   0·001). Platelet VEGF reflected the reactivity of IVIG treatment and was decreased in responders ( P  <   0·001), but remained increased in non-responders ( P  =   0·01). Platelet VEGF levels, but not serum VEGF levels, before IVIG were significantly correlated with the maximum CAA z -score ( r  = 0·524, P  =   0·02). Our findings demonstrate that platelet VEGF may reflect the severity of vasculitis related to the pathological development of CAA in KS. Platelet VEGF may be an important feature of KS pathophysiology.     

Ultrasound of peripheral nerves in acromegaly: changes at 1-year follow-up

Context  We have previously demonstrated peripheral nerve enlargement in acromegaly.
Objective  The aim of this study was to use ultrasound (US) to assess any changes in the peripheral nerves of patients with acromegaly 1 year after the first evaluation.
Patients  We prospectively examined the median and ulnar nerve cross-sectional area (CSA) in 34 non-diabetic, patients with acromegaly (18 females and 16 males; 18–79 years) and 34 age-, sex-, BMI-matched controls, using a 17–5 MHz US probe.
Intervention  The median nerve was examined at the mid-forearm (MN-f) and at the carpal tunnel (MN-Ct) levels; the ulnar nerve at mid-forearm (UN-f) and at distal arm (UN-a). Patients were grouped according to the clinical control of the disease: 'improved'; 'always controlled'; 'always uncontrolled'; and 'worsened'.
Results  The median nerve at mid-forearm (MN-f), the ulnar nerve at mid-forearm (UN-f) and at distal arm (UN-a) were significantly reduced after 1-year follow-up in all patients ( P <  0·001, P  < 0·008, P  < 0·012, respectively). In the 'improved' group, there was a significant reduction of median nerve CSA examined at mid-forearm (MN-f) ( P =  0·02), and distal arm ulnar nerve CSA (UN-a) ( P =  0·002). In the other groups no statistically significant differences in ultrasound parameters were recorded. However, UN-a, UN-f, MN-f, MN-ct were still significantly higher in all groups compared with controls ( P <  0·001).
Conclusion  These data demonstrate that median and ulnar nerves CSA are reduced after 1 year follow-up, in line with the reduction of GH/IGF-I levels. However, as the control of the disease incompletely reverts nerve enlargement, this phenomenon could be only partially reversible.     

Primary tumour diameter as a risk factor for advanced disease features of differentiated thyroid carcinoma

Objective  To study the relationship between primary tumour size and the risk of advanced disease features (multifocal or locally invasive disease, lymph-node or distant metastases) in differentiated thyroid carcinoma (DTC).
Design  A retrospective chart review study.
Patients  The study sample comprised 935 papillary (PTC) and 291 follicular thyroid carcinoma (FTC) patients treated in our hospital from 1978 to 2007.
Measurements  Kaplan–Meier analyses and log-rank tests were performed to calculate tumour size-adjusted cumulative risk of advanced disease features.
Results  Accounting for primary tumour diameter, there were no significant differences in cumulative risks of multifocal carcinoma ( P =  0·12) or distant metastases ( P =  0·49) between PTC and FTC. PTC showed higher cumulative risks of local invasion ( P <  0·0001) or lymph-node metastases ( P <  0·0001). The cumulative risk of tumour multifocality increased 5%/cm of primary tumour diameter. The cumulative risk of local invasion or lymph-node metastases in PTC and of distant metastases in DTC increased exponentially at a threshold tumour diameter of 10 mm. In FTC, lymph-node metastases are associated almost exclusively with primary tumours showing extrathyroidal growth.
Conclusions  Starting with a 1 cm primary tumour diameter, increasing tumour size is associated with an exponentially increasing risk of local invasion or lymph-node or distant metastases of DTC. The current classification of carcinomas < 2 cm as T1 is therefore questionable.     

Bone marrow microvessel density in chronic myeloproliferative disorders: a study of 115 patients with clinicopathological and molecular correlations

Philadelphia-negative chronic myeloproliferative disorders (CMD) include polycythemia vera (PV), essential thrombocythemia (ET) and primary myelofibrosis (PMF). Angiogenesis is critical in the pathogenesis of PMF. We studied angiogenesis in 115 patients with CMD (23 PV, 24 ET, 46 PMF, 12 post-PV and 10 post-ET myelofibrosis) by assessment of microvessel density (MVD) in bone marrow (BM). Kruskall–Wallis analysis of variance showed that patients with PMF had significantly higher values of MVD than those with PV ( P  <   0·001), ET ( P  <   0·001) and controls ( P  <   0·001). Mann–Whitney U -test demonstrated that patients with PMF at the prefibrotic stage had significantly higher MVD values than those with ET ( P  =   0·02). Patients with post-PV myelofibrosis showed significantly higher MVD values than those with PV ( P  <   0·001), as did patients with post-ET myelofibrosis compared with ET ( P  <   0·001). In patients with CMD, the multivariate generalized linear regression model showed that the JAK2 (V617F) mutational burden ( P  =   0·01), serum lactate dehydrogenase level ( P  =   0·003), and anaemia ( P  <   0·001) independently correlated with MVD. In summary, this study indicates that assessment of BM angiogenesis, as measured by MVD, may be a useful additional tool in the histopathological definition of CMD.     

A virally inactivated functional growth factor preparation from human platelet concentrates

Background  Human platelet growth factors (HPGF) are essential for tissue regeneration and may replace fetal bovine serum (FBS) in cell therapy. No method for the manufacture of standardized virally inactivated HPGF has been developed yet.
Study design and methods  Platelet concentrates (PC) were subjected to solvent/detergent (S/D) treatment (1% TnBP/1% Triton X-45), oil extraction, hydrophobic interaction chromatography and sterile filtration. Platelet-derived growth factor (PDGF)-AB, -BB and -AA, transforming growth factor-ß1 (TGF-ß1), epidermal growth factor (EGF), insulin-like growth factor-1 (IGF-1) and vascular endothelium growth factor (VEGF) were measured by ELISA. Composition in proteins and lipids was determined, protein profiles were obtained by SDS-PAGE, and TnBP and Triton X-45 were assessed by gas chromatography and high-performance liquid chromatography, respectively. Cell growth promoting activity of HPGF was evaluated by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay using human embryonic kidney (HEK293A) fibroblast and Statens Seruminstitute rabbit corneal (SIRC) epithelial cell lines.
Results  The GF preparation contained a mean of 16·66, 2·04, 1·53, 72·19, 0·33, 48·59 and 0·44 ng/ml of PDGF-AB, -BB, -AA, TGF-ß1, EGF, IGF-1 and VEGF, respectively. The protein profile was typical of platelet releasates and had less than 2 p.p.m. of residual S/D agents. MTS assay of HEK293A and SIRC cultures showed that the GF preparation at 10% and 0·1% (v/v), respectively, could successfully replace 10% FBS for cell proliferation. Cell-stimulating activity of HPGF on HEK293A was over twice that of PC releasates.
Conclusion  Standardized and functional virally inactivated HPGF can be prepared from human PC for possible applications in cell therapy and regenerative medicine.     

Higher serum TSH in thyroid cancer patients occurs independent of age and correlates with extrathyroidal extension

Background  It has previously been shown that higher serum TSH is associated with increased thyroid cancer incidence and advanced-stage disease. In the healthy adult population, mean TSH increases with age. As age over 45 years is a known prognostic indicator for thyroid cancer, it is important to know whether higher TSH in patients with thyroid cancer occurs independent of age.
Objective  To determine the relationship between higher TSH, cancer and age.
Design  A retrospective cohort study.
Patients and methods  A total of 1361 patients underwent thyroid surgery between May 1994 and December 2007 at a single institution. Of these patients, 954 had pathological data, preoperative TSH and complete surgical history available. Data were analysed in relation to age and TSH.
Results  Mean TSH was significantly higher in cancer patients regardless of age < 45 years or ≥ 45 years ( P =  0·046 and P  = 0·027, respectively). When examining age groups < 20, 20–44, 45–59 and ≥ 60 years, there was a trend of rising mean TSH with age. Despite the rise in the benign subgroups, mean TSH was consistently higher in those with cancer vs. those without. On multivariate analysis, higher TSH was independently associated with cancer ( P =  0·039) and pathological features of Hashimoto's thyroiditis ( P =  0·001) but not with age ( P =  0·557). On multivariate analysis of high-risk features associated with poor prognosis, there was a significant association between higher TSH and extrathyroidal extension ( P =  0·002), whereas there was no clear relationship with age, tumour size > 4 cm, and distant metastases.
Conclusion  Independent of age, thyroid cancer incidence correlates with higher TSH. Higher TSH is associated with extrathyroidal extension of disease.     

共同培养诱导骨髓基质干细胞向肝细胞分化的研究

目的 探索大鼠骨髓皋质干细胞（MsCS）向肝细胞分化的能力，及肝细胞生长的微环境埘其诱导分化的作用。方法 采用梯度离心法，获取大鼠骨髓基质干细胞；改良的两步法获取大鼠肝细胞。将鉴定的MsCS和肝细胞以半透膜相隔共同培养，以单独培养的MSCs作对照。在第1、3、7．14．21、28天，分别以逆转录聚合酶链反应（RT-PCR）和免疫细胞化学分析检测甲胎蛋白（AFP）、白蛋白、细胞角蛋白l8（CK-l8）的基因和蛋白表达。结果在MSCs与肝细胞共同培养过程中，MSCs出现明显的细胞形态、体积和数量变化，可见双核或多核细胞，细胞轮廓较清晰。RT-PCR检测：共同培养的MSCs第7天即出现AFP基因表达，第14天表达增强，第21天表达减弱；第14天开始出现白蛋白、CK-18基因表达，并持续表达。单独培养的MSCs均无表达。共同培养的MSCs，于第7天进行免疫细胞化学检测，AFP即呈阳性；第14天白蛋白和CK-18也呈阳性；单独培养的MSCs未见AFP、白蛋白及CK-18表达。结论 大鼠骨髓基质干细胞与肝细胞共同培养，可被诱导分化为肝细胞。     

In vitro differentiation of mouse bone marrow mononuclear cells into hepatocyte-like cells.

AIM:: It is imperative to explore some ways to gain the functional hepatocytes for hepatocyte transplantation. Bone marrow stem cells can differentiate into hepatocytes in vivo and in vitro. We select fibroblast growth factor-4 (FGF-4), oncostatin M (OSM), hepatocyte growth factor (HGF) and epithermal growth factor (EGF) as differentiation factors, and design the appropriate directed differentiation medium in order to gain hepatocytes through directed differentiation of bone marrow stem cells. METHODS:: Bone marrow mononuclear cells (BMMCs) were cultured in the directed differentiation media including FGF-4, OSM, HGF and EGF. In the course of cell differentiation, cell morphology was observed, and the expression patterns of some genes of the hepatocyte were validated and confirmed by RT-PCR. The ALB-, and CK18-expressed cells were gone further step to be confirmed by Western blot analysis, immunofluorescence and flow cytometric analysis. Hepatocyte functional activity, including glycogen synthesis and urea production, were confirmed by periodic acid-Shiff (PAS) staining and urea assay. RESULTS:: Some epithelial-like cells or polygonal cells appeared in the directed differentiation medium within 12 days, and the number and sizes of colonies of epithelial-like cells or polygonal cells increased in the course of the cell directed differentiation. AFP, HNF-3ss, ALB and CK18 mRNA expressions first appeared within day 7, and lasted throughout the later directed differentiation. TTR, G-6-P and TAT mRNA expressions could be detected within day 14, and their expressions lasted in the course of the later directed differentiation. ALB and CK18 were confirmed to exist in the differentiated BMMCs by Western blot analysis. ALB was found in the cytoplasm and cell membrane, while CK18 scattered in the cytoplasm by immunofluorescent staining. On day 21,the ratio of ALB-positive cells was 69.45%, and the ratio of CK18-positive cells was 67.36%. The accumulation of glucogen was detected in the cytoplasm of the differentiated cells. The directed differentiated BMMCs produced urea 3 days later, and they produced urea in a time-dependent manner. CONCLUSIONS:: BMMCs could differentiate into hepatocytes or hepatocyte-like cells in the differentiation media including HGF, FGF-4, EGF, and OSM. These hepatocyte-like cells were identified at the gene level and protein level. Furthermore, these hepatocyte-like cells had some hepatocellular synthesis and metabolism functions.     

Peroxisome proliferator-activated receptor gamma agonism modifies the effects of growth hormone on lipolysis and insulin sensitivity

Context  Peroxisome proliferator-activated receptor gamma (PPAR-γ) agonists such as thiazolidinediones (TZDs) improve insulin sensitivity in type 2 diabetes mellitus (T2DM) through effects on fat metabolism whereas GH stimulates lipolysis and induces insulin resistance.
Objective  To evaluate the impact of TZDs on fat metabolism and insulin sensitivity in subjects exposed to stable GH levels.
Design  A randomized, placebo-controlled, double-blind parallel-group study including 20 GH-deficient patients on continued GH replacement therapy. The patients were studied before and after 12 weeks.
Intervention  Patients received either pioglitazone 30 mg ( N  = 10) or placebo ( N  = 10) once daily for 12 weeks.
Results  Adiponectin levels almost doubled during pioglitazone treatment ( P =  0·0001). Pioglitazone significantly decreased basal free fatty acid (FFA) levels ( P =  0·02) and lipid oxidation ( P =  0·02). Basal glucose oxidation rate ( P =  0·004) and insulin sensitivity ( P =  0·03) improved in the patients who received pioglitazone treatment. The change in insulin-stimulated adiponectin level after pioglitazone treatment was positively correlated to the change in insulin-stimulated total glucose disposal ( R  = 0·69, P  = 0·04).
Conclusion  The impact of GH on lipolysis and insulin sensitivity can be modified by administration of TZDs.     

Efficacy of transfusion of platelet concentrates obtained by manual pooling or by semiautomated pooling of buffy-coats: a retrospective analysis of count increment, corrected count increment and transfusion interval

Background and Objective  The preparation of platelet concentrates from pooling buffy-coats can be done manually or semiautomatically with the OrbiSac BC System. The objective of this study was to compare the clinical outcome of platelet transfusions prepared by these two methods in terms of 1-h count increment (1-h CI), 1-h corrected count increment (1-h CCI) and transfusion interval.
Materials and Methods  Platelet transfusions were identified retrospectively for inclusion in the control group (manual pooling) or the test group (automated pooling). Transfusion outcome variables were 1-h CI, 1-h CCI and transfusion interval. The impact of 16 patient- and platelet concentrate-related variables on transfusion efficacy was investigated by multiple regression analysis.
Results  The database analysed consisted of 205 control transfusions given to 36 patients and 219 test transfusions given to 36 patients. Mean platelet content and mean 1-h CI were statistically higher in the test group when compared to the control group ( P <  0·05), but mean 1-h CCI and transfusion interval were not. Multiple regression analysis resulted in models explaining 13% of the variance of 1-h CI, 8% of the variance of 1-h CCI and 17% of the variance of transfusion interval ( P <  0·05).
Conclusion  There are no significant differences between the two preparation methods regarding the clinical outcome of platelet transfusions, as the difference in 1-h CI is explained by differences in platelet content.     

The presence of CD56/CD16 in T-cell acute lymphoblastic leukaemia correlates with the expression of cytotoxic molecules and is associated with worse response to treatment

Some cases of T-cell acute lymphoblastic leukaemia (ALL) express markers found in natural-killer (NK) cells, such as CD56 and CD16. Out of 84 T-cell ALL cases diagnosed at our Institution, CD56 and/or CD16 was detected in 24 (28·5%), which we designated T/NK-ALL group. Clinical features, laboratory characteristics, survival and expression of cytotoxic molecules were compared in T/NK-ALL and T-ALL patients. Significant differences were observed regarding age (24·9 vs. 16·4 years in T/NK-ALL and T-ALL, respectively, P  =   0·006) and platelet counts (177 × 10⁹/l vs. 75 × 10⁹/l in T/NK-ALL and T-ALL, respectively, P  =   0·03). Immunophenotypic analysis demonstrated that CD34, CD45RA and CD33 were more expressed in T/NK-ALL patients, whereas CD8 and terminal deoxynucleotidyl transferase were more expressed in T-ALL patients ( P  <   0·05 ) . The mean overall survival (863 vs. 1869 d, P  =   0·02) and disease-free survival (855 vs. 2095 d, P  =   0·002) were shorter in patients expressing CD56/CD16. However, multivariate analysis identified CD56/CD16 as an independent prognostic factor only for DFS. Cytotoxic molecules were highly expressed in T/NK-ALL compared to T-ALL. Perforin, granzyme B and TIA-1 were detected in 12/17, 4/17 and 7/24 T/NK-ALL patients and in 1/20, 0/20 and 1/20 T-ALL respectively ( P  <   0·001, P  =   0·036 and P  =   0·054). Therefore, the presence of CD56/CD16 was associated with distinct clinical features and expression of cytotoxic molecules in the blasts.     

Decreased hepatic RBP4 secretion is correlated with reduced hepatic glucose production but is not associated with insulin resistance in patients with liver cirrhosis

Objective  Patients with liver cirrhosis have a high incidence of insulin resistance and diabetes. This study was designed to determine circulating levels and hepatic production of retinol-binding protein 4 (RBP4) in relation to parameters of hepatic and systemic metabolism in patients with liver cirrhosis.
Design and method  Circulating RBP4 levels were measured in 19 patients with liver cirrhosis at different clinical stages of the disease and in 20 age-, sex- and body mass index (BMI)-matched controls. Hepatic production rates of RBP4 and glucose were assessed by measuring the arterial hepatic venous concentration difference together with hepatic blood flow. Insulin resistance was determined by the Quantitative Insulin Sensitivity Check Index (QUICKI) and the homeostasis model assessment of insulin resistance (HOMA-IR), energy expenditure by indirect calorimetry and body composition by bioelectrical impedance analysis (BIA).
Results  Compared with controls, RBP4 levels in cirrhosis were decreased (8·1 ± 1·8 vs. 22·6 ± 2·4 mg/l, P  < 0·001) due to decreased hepatic production ( P <  0·05). RBP4 correlated with hepatic protein synthesis capacity ( P <  0·01), but not with insulin resistance, energy expenditure, BMI or body fat mass. Plasma RBP4 correlated with hepatic glucose production ( P <  0·05).
Conclusions  These data demonstrate that RBP4 in cirrhosis (i) is decreased due to reduced hepatic production, (ii) is not associated with insulin resistance, and (iii) might have a beneficial role by decreasing hepatic glucose production and could thus also be regarded as a hepatokine.     

Efficient generation of functional hepatocytelike cells from mouse liver progenitor cells via indirect co-culture with immortalized human hepatic stellate cells

BACKGROUND: Differentiation of liver progenitor cells(LPCs) to functional hepatocytes holds great potential to develop new strategies for hepatocyte transplantation and the screening of drug-induced cytotoxicity. However, reports on the efficient and convenient hepatic differentiation of LPCs to hepatocytes are few. The present study aims to investigate the possibility of generating functional hepatocytes from LPCs in an indirect co-culture system.METHODS: Mouse LPCs were co-cultured in Transwell plates with an immortalized human hepatic stellate cell line(HSCLi) we previously established. The morphology, expression of hepatic markers, and functions of mouse LPC-derived cells were monitored and compared with those of conventionally cultured LPCs. RESULTS: Co-culturing with HSC-Li cells induced differentiation of mouse LPCs into functional hepatocyte-like cells. The differentiated cells were morphologically transformed into hepatocyte-like cells 3 days after co-culture initiation. In addition, the differentiated cells expressed liver-specific genes and possessed hepatic functions, including glycogen storage, lowdensity lipoprotein uptake, albumin secretion, urea synthesis, and cytochrome P450 1A2 enzymatic activity.CONCLUSIONS: Our method, which employs indirect co-culture with HSC-Li cells, can efficiently induce the differentiation of LPCs into functional hepatocytes. This finding suggests that this co-culture system can be a useful method for the efficient generation of functional hepatocytes from LPCs.