Culture of 3D bioprinted bone constructs requires an increased fluid dynamic stimulation.

In vitro flow-induced mechanical stimulation of developing bone tissue constructs has been shown to favor mineral deposition in scaffolds seeded with cells directly exposed to the fluid flow. However, the effect of fluid dynamic parameters, such as shear stress (SS), within 3D bioprinted constructs is still unclear. Thus, this study aimed at correlating the SS levels and the mineral deposition in 3D bioprinted constructs, evaluating the possible dampening effect of the hydrogel. Human mesenchymal stem cells (hMSCs) were embedded in 3D bioprinted porous structures made of alginate and gelatin. 3D bioprinted constructs were cultured in an osteogenic medium assessing the influence of different flow rates (0, 0.7 and 7 ml/min) on calcium and collagen deposition through histology, and bone volume (BV) through micro-computed tomography. Uniform distribution of calcium and collagen was observed in all groups. Nevertheless, BV significantly increased in perfused groups as compared to static control, ranging from 0.35±0.28 mm3, 11.90±8.74 mm3 and 25.81±5.02 mm3 at week 3 to 2.28±0.78 mm3, 22.55±2.45 mm3 and 46.05±5.95 mm3 at week 6 in static, 0.7 and 7 ml/min groups, respectively. SS values on construct fibers in the range 10-100 mPa in 7 ml/min samples were twice as high as those in 0.7 ml/min samples showing the same trend of BV. The obtained results suggest that it is necessary to enhance the flow-induced mechanical stimulation of cell-embedding hydrogels to increase the amount of mineral deposited by hMSCs, compared to what is generally reported for the development of in vitro bone constructs. STATEMENT OF SIGNIFICANCE: In this study, we evaluated for the first time how the hydrogel structure dampens the effect of flow-induced mechanical stimulation during the culture of 3D bioprinted bone tissue constructs. By combining computational and experimental techniques we demonstrated that those shear stress thresholds generally considered for culturing cells seeded on scaffold surface, are no longer applicable when cells are embedded in 3D bioprinted constructs. Significantly, more bone volume was formed in constructs exposed to shear stress values generally considered as detrimental than in constructs exposed shear stress values generally considered as beneficial after 3 weeks and 6 weeks of dynamic culture using a perfusion bioreactor.

Mortality Analysis of Acute Liver Failure in Uruguay.

BACKGROUND: Acute liver failure (ALF) is a syndrome with high mortality. OBJECTIVE: Describe characteristics and outcomes of patients with ALF in Uruguay, and identify factors associated with mortality. METHODS: A retrospective analysis of 33 patients with ALF was performed between 2009 and 2017. RESULTS: The patients' median age was 43 years, and 64% were women. Average Model for End-Stage Liver Disease (MELD) score at admission was 33. The median referral time to the liver transplant (LT) center was 7 days. The most common etiologies were viral hepatitis (27%), indeterminate (21%), autoimmune (18%), and Wilson disease (15%). Overall mortality was 52% (71% of transplanted and 46% of nontransplanted patients). Dead patients had higher referral time (10 vs 4 days, P = .008), higher MELD scores at admission (37 vs 28) and highest achieved MELD scores (42 vs 29; P < .001), and higher encephalopathy grade III to IV (94% vs 25%, P < .001) than survivors. Patients without LT criteria (n = 4) had lower MELD score at admission (25 vs 34, P = .001) and highest achieved MELD score (27 vs 37, P = .008) compared with the others. Patients with LT criteria but contraindications (n = 7) had higher MELD scores at admission (38 vs 31, P = .02), highest achieved MELD scores (41 vs 34, P = .03), and longer referral time (10 days) than those without contraindications (3.5 days) or those without LT criteria (7.5 days, P = .02). Twenty-two patients were listed; LT was performed in 7, with a median time on waiting list of 6 days. CONCLUSIONS: ALF in Uruguay has high mortality associated with delayed referral to the LT center, MELD score, and encephalopathy. The long waiting times to transplantation might influence mortality.

Risk Factors of Mortality After Liver Transplantation in Uruguay.

INTRODUCTION: Identification of predictive factors of mortality in a liver transplant (LT) program optimizes patient selection and allocation of organs. OBJECTIVE: To determine survival rates and predictive factors of mortality after LT in the National Liver Transplant Program of Uruguay. METHODS: A retrospective study was conducted analyzing data prospectively collected into a multidisciplinary database. All patients transplanted since the beginning of the program on July 2009 to April 2017 were included (n = 148). Twenty-nine factors were analyzed through the univariate Kaplan-Meier model. A Cox regression model was used in the multivariate analysis to identify the independent prognostic factors for survival. RESULTS: Overall survival was 92%, 87%, and 78% at discharge, 1 year, and 3 years, respectively. The Kaplan-Meier survival curves were significantly lower in: recipients aged >60 years, Model for End-Stage Liver Disease score >21, LT due to hepatocellular carcinoma (HCC) and acute liver failure (ALF), donors with comorbidities, intraoperative blood loss beyond the median (>2350 mL), red blood cell transfusion requirement beyond the median (>1254 mL), intraoperative complications, delay of extubation, invasive bacterial, and fungal infection after LT and stay in critical care unit >4 days. The Cox regression model (likelihood ratio test, P = 1.976 e-06) identified the following independent prognostic factors for survival: LT for HCC (hazard ratio [HR] 4.511; P = .001) and ALF (HR 6.346; P = .004), donors with comorbidities (HR 2.354; P = .041), intraoperative complications (HR 2.707; P = .027), and invasive fungal infections (HR 3.281; P = .025). CONCLUSION: The survival rates of LT patients as well as the mortality-associated factors are similar to those reported in the international literature.

Isolation and characterization of an antitumor active agar-type polysaccharide of Gracilaria dominguensis.

Cold water extraction of the red alga Gracilaria dominguensis, followed by cetyltrimethylammonium bromide fractionation, gave a highly sulfated, agar-type polysaccharide which inhibited the transplantation of Ehrlich ascites carcinoma in mice. The structure of the polysaccharide has been investigated by methylation analysis, and 1H- and 13C-n.m.r. spectroscopy, and was shown to be mainly composed of alternating (1----3)-linked beta-D-galactopyranosyl 6-sulfate and (1----4)-linked 3,6-anhydro-alpha-L-galactopyranosyl residues.

Effect of two antiproteinases on the growth of transplantable tumors and the proliferation of untransformed and transformed cells in culture.

The paper reports on the experiments carried out for the evaluation of the effect of two antiproteinases, aprotinin, and epsilon-amino caproic acid (EACA), on several transplantable tumors and cells in culture. It was demonstrated that aprotinin has a preferential therapeutic effect on the solid form of L1210, in comparison with its ascitic form. Treatment with aprotinin of Lewis lung carcinoma, Melanoma B16, and Hepatoma 22, in a limited chronic time schedule 1-9, and 1-11 brought no significant increase in survival; positive therapeutic effects had been shown for the first and the last of the tumors mentioned with life-time injections of the drug. Moreover, aprotinin treatment increased the number of lung nodules for the Lewis lung carcinoma inoculated either subcutaneously or intravenously. EACA applied same schedule had no effect on the survival of tumor-bearing animals. These data are discussed in terms of their relevance to antiproteinase therapy in human cancer. No selective inhibition of proliferation for more tumorigenic cell culture lines of spontaneous and viral origin was demonstrated after treatment with both compounds.

[Immunodeficiencies and enzyme deficiencies]. / Immunodeficienze e deficit enzimatici.

The first part of the paper outlines the actual situation of the recent investigations on the interpretative problem of the immunological deficits combined with enzymatic deficits. ADA deficit associated with combined immunodeficiency is an autosomal recessive form. Low enzyme levels, which is produced by a structural gene located on chromosome 20, were detected in chronic lymphatic leukemias where ADA decrease is correlated to low levels of T lymphocytes and the prevalence of an atypical B lymphocyte population. Particularly high levels of ADA were detected in acute lymphoblastic leukemias, in transplant rejection, in blastic crisis of chronic myeloid leukemia... NP deficit is associated with a T branch immunodeficiency, with high levels of inosine, guanosine, hypouricemia, hypochromic microcythemia and hematopoietic tissue megaloblastosis. This enzyme, with trimeric structure, whose structural gene is located on chromosome 14, shows a cytoplasmic location, and its maximum activity is to be found in T lymphocytes separated by rosetting. IGPT deficit is to be held responsible for the neurological Lesch-Nyhan syndrome. This deficit is associated with a depression of B lymphocyte function evaluated as response to the mitogens (PWM, Protein A) or as specific immunoglobulins production. At last the Authors report some personal investigations performed on hemolysates and lymphocytes of subjects with impaired immunity, as well as on some children at birth to establish the correlation between ADA and NP behaviour and immunosurveillance. Lastly, the data on the variations of the enzymatic and immunological parameters of subjects with immunodeficiency associated to enzymopenia after red cell transfusion are reported.