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Biomedical subjects

Koichi Tanaka

Publications and source records attributed to Koichi Tanaka.

At least 91 records · Page 5Linked to original sources
PubMed2005

Living-donor liver transplantation for hepatoblastoma.

Hepatoblastoma is the most common malignant liver tumor in children. Recently, liver transplantation has been indicated for unresectable hepatoblastoma. We retrospectively reviewed 14 children with a diagnosis of hepatoblastoma who had undergone living-donor liver transplantation (LDLT) at Kyoto University Hospital. During the period from June 1990 to December 2004, 607 children underwent LDLT. Of these interventions, 2.3% were performed for hepatoblastoma. Based on radiological findings, the pre-treatment extent of disease (PRETEXT) grouping was used for pre-treatment staging of the tumor. There were grade III in seven patients and grade IV in seven patients. Thirteen patients received chemotherapy, and seven underwent hepatectomy 11 times. Immunosuppressive treatment consisted of tacrolimus monotherapy in 11 patients. Actuarial 1- and 5-year graft and patient survival rates were 78.6% and 65.5%. The poor prognostic factors were macroscopic venous invasion and extrahepatic involvement with 1-year and 5-year survival rates of 33.0% and 0%. Pediatric patients without these factors showed an acceptable 5-year survival rate of 90.9%. LDLT provides a valuable alternative with excellent results in children with hepatoblastoma because it allows optimal timing of the liver transplantation, given the absence of delay between the completion of chemotherapy and planned liver transplantation.

Chemotherapy, Adjuvant↗
PubMed2005

Living donor liver transplantation for pediatric patients with inheritable metabolic disorders.

Forty-six pediatric patients who underwent living donor liver transplantation (LDLT) using parental liver grafts for inheritable metabolic disorders (IMD) were evaluated to determine the outcomes of the surgery, decisive factors for post-transplant patient survival and the impact of using donors who were heterozygous for the particular disorder. Disorders included Wilson disease (WD, n = 21), ornithine transcarbamylase deficiency (OTCD, n = 6), tyrosinemia type I (TTI, n = 6), glycogen storage disease (GSD, n = 4), propionic acidemia (PPA, n = 3), methylmalonic acidemia (MMA, n = 2), Crigler-Najjar syndrome type I (CNSI, n = 2), bile acid synthetic defect (BASD, n = 1) and erythropoietic protoporphyria (EPP, n = 1). The post-transplant cumulative patient survival rates were 86.8 and 81.2% at 1 and 5 years, respectively. Post-transplant patient survival and recovery of the growth retardation were significantly better in the liver-oriented diseases (WD, OTCD, TTI, CNSI and BASD) than in the non-liver-oriented diseases (GSD, PPA, MMA and EPP) and pre-transplant growth retardation disadvantageously affected post-transplant outcomes. Although 40 of 46 donors were considered heterozygous for each disorder, neither mortality nor morbidity related to the heterozygosis has been observed. LDLT using parental donors can be recommended as an effective treatment for pediatric patients with IMD. In the non-liver-oriented diseases, however, satisfactory outcomes were not obtained by hepatic replacement alone.

Adolescent↗
PubMed2005

Therapeutic strategy and the role of apheresis therapy for ABO incompatible living donor liver transplantation.

Although in Japan, transplant organs obtained from brain-dead donors (BDD) has been allowed since October 1997, to date only 27 liver grafts from BDD have been obtained. The severe shortage of transplantable organs is a big problem, not only in liver transplantation but also other organ transplants. Liver transplantation is a fundamental treatment for end-stage liver disease. In order to perform living-donor liver transplantation (LDLT) in a safer manner, apheresis (plasmapheresis) plays a major role in Japan because of the prevalence of LDLT wherein later re-transplantation is difficult. Therefore, because of a limited donor supply and because the need of patients with end-stage liver disease is critical, use of grafts from ABO-incompatible donors may be the only available option. From June 1990 to November 2004, 1010 patients underwent 1060 LDLT cases at Kyoto University Hospital. Of these, 139 LDLT cases (13.1%) received ABO-incompatible living-donor liver grafts. The role of apheresis in ABO-incompatible LDLT is the reduction of antibody titers such as anti-A or anti-B antibody. We perform preoperative apheresis as a general rule for incompatible cases, and the recipient's antibody level against the donor's blood type is decreased to one eighth of the baseline value before LDLT. Up to the present, baseline antirejection regimens included steroids, tacrolimus and cyclophosphamide. At first, splenectomy was performed during operation to suppress antibody production, and intraportal infusion therapy was performed to control local disseminated intravascular coagulation (DIC) occurring in ABO-incompatible grafts. At that time, three agents--methylprednisolone, prostaglandin E1, and gabexate mesilate--were infused continuously for 3 weeks after LDLT. At present, instead of intraportal infusion therapy, hepatic artery infusion therapy without splenectomy is adopted because of portal thrombosis, and two agents--methylprednisolone and prostaglandin E1--are infused continuously for 3 weeks after LDLT. Recently, we introduced an anti-CD20 monoclonal antibody (Rituximab) instead of splenectomy for B cell deletion before ABO-incompatible LDLT. In this article, we describe our therapeutic strategy and the role of apheresis around ABO-incompatible LDLT.

ABO Blood-Group System↗
PubMed2005

Localization of aquaporin water channels in the airway of the musk shrew (Suncus murinus) and the rat.

Aquaporins (AQPs) constitute a family of water channels that facilitate membrane water permeability in various tissues of animals. In this study, we compared the expression and localization of AQPs in the respiratory system of the musk shrew (Suncus murinus), which is an insectivore, and the rat by immunohistochemical methods. In both the musk shrew and the rat, AQP1 was expressed throughout the airway in endothelial cells of subepithelial blood vessels and in nasal submucosal fibroblasts. AQP3 and AQP4 were detected in neither the epithelium nor the subepithelial layer of the musk shrew airway, but were abundant in the rat airway epithelium. Musk shrew AQP5 was distributed in the superficial epithelial cells facing the airspaces and in submucosal glandular cells, but, unlike in the rat, not in lung alveolar cells. Additionally, the expression patterns of AQP4 and AQP5 of the musk shrew were partly similar to those of the human previously reported, absence of AQP4 and presence of AQP5 in the upper airway. The expression differences of AQPs between species in the airway indicate that the physiological importance of each AQP may be different in each species.

Animals↗
PubMed2005

Effective islet isolation method with extremely high islet yields from adult pigs.

Achieving good islet isolation is one of the most important factors for successful islet transplantation. Porcine pancreas is suitable for islet isolation research due to its anatomical and physiological similarities to human pancreas. In this study, we evaluated a new porcine islet isolation method designed to maximize islet yield and compared it with our previous open pan method and the standard method using a Ricordi chamber (Ricordi method). We performed 15 porcine islet isolations, five each with the new method, the open pan method, and the Ricordi method. The new method features several important improvements. Pancreata remain uncut and are kept intact during collagenase intraductal injection, a large filtration chamber to handle whole pancreata, low concentration of collagenase (Liberase HI) for digestion, and large plastic containers for large-scale islet purification. All isolated islets were assessed for yield, purity, viability and in vitro function. Islets isolated with this new method were transplanted under the kidney capsules of SCID mice with chemically induced diabetes for in vivo functional assessment (n = 8). With the new method, we obtained on average more than 1,000,000 islet equivalents (IE) (1,236,266 +/- 213,486 IE) (mean +/- SE) before purification and 800,000 IE (879,815 +/- 222,729 IE) after purification from one adult pig. Islet yield per pancreas was significantly higher compared with our previous open pan method (30,666 +/- 11,532 IE, p < 0.01) and the Ricordi method (317,073 +/- 86,093 IE, p < 0.05). All mice, transplanted with 1000 islets from the new method, returned to normoglycemia within 4 days after transplantation. Our new method makes it possible to obtain extremely high porcine islet yield with good function. It should produce useful information for human islet isolation and transplantation, and might be applied to single donor clinical xenogeneic transplantation.

Animals↗
PubMed2005

PDX-1 protein is internalized by lipid raft-dependent macropinocytosis.

PDX-1 plays a central role in regulating insulin gene transcription and differentiation of insulin-producing cells. It was previously reported that, due to its own Antennapedia-like protein transduction domain (PTD), exogenous PDX-1 protein can permeate cells and induces insulin gene expression in pancreatic ducts, thought to be islet progenitor cells. These data suggest that PDX-1 protein transduction could be a safe and valuable strategy for facilitating differentiation of progenitor cells into insulin-producing cells without requiring gene transfer technology. Here it is shown that after an initial ionic cell-surface interaction, PDX-1 proteins are rapidly internalized by lipid raft-dependent macropinocytosis. HeLa cells were treated with both FITC-conjugated PDX-1 PTD and FM 4-64, a general fluorescent marker of endocytosis. A punctate cytoplasmic distribution of PDX-1 PTD, which colocalized with FM 4-64, was observed in treated cells. Because expression of dominant-negative dynamin-1 did not block PDX-1 PTD uptake, PDX-1 protein transduction is independent on phagocytosis and clathrin- or caveolar-mediated endocytosis. Cells were pretreated with amiloride, a specific inhibitor of the Na+/H+ exchange required for macropinocytosis, or cytochalasin D, an F-actin elongation inhibitor. Treatment of cells with both macropinosome inhibitors resulted in the reduction in PDX-1 PTD transduction into vesicles, suggesting that PDX-1 PTD-mediated cellular entry occurs by lipid raft-mediated macropinocytosis. Taken together, these observations provide the mechanism of PDX-1 protein transduction and suggest that the protein transduction system could work for experimental and therapeutic strategies.

Biological Transport↗
PubMed2005

Pathologic significance of low-attenuation hepatic parenchymal abnormalities in CT scans of living related donor partial liver transplant recipients.

PURPOSE: To evaluate the pathologic significance of the low-attenuation changes of liver grafts in living donor partial liver transplantation (LDLT) on computed tomography (CT). MATERIALS AND METHODS: We retrospectively correlated the low-attenuation changes of liver grafts which were defined as readings below 50 HU on unenhanced CT with histological findings obtained by needle biopsy or surgery within seven days of CT studies. The study group included 35 CT findings of 35 recipients. We classified the low-attenuation change of the liver grafts into a homogeneous low-attenuation group and a heterogeneous low-attenuation group. RESULTS: Major histologic findings of 35 specimens included cholangitis in 10 (28.6%), cholestasis in eight (22.9%), fatty change in 15 (42.9%), acute cellular rejection (ACR) in 12 (34.3%), acute hepatitis in three (8.6%), liver cell ballooning in four (11.4%), massive hepatic necrosis in four (11.4%), and centrilobular congestion in five (14.3%) cases, respectively. Homogeneous and heterogeneous low-attenuation changes of liver grafts on unenhanced CT were seen in 26 (74.3%) and 9 (25.7%) cases, respectively. Massive hepatic necrosis occurred more frequently among the heterogeneous low-attenuation group than among the homogeneous low-attenuation group. CONCLUSION: Low-attenuation changes in liver grafts on CT may indicate a variety of pathological changes. Heterogeneous low-attenuation changes suggest massive hepatic necrosis.

Adolescent↗
PubMed2004

"Small-for-size graft" and "small-for-size syndrome" in living donor liver transplantation.

The indications for living donor liver transplantation (LDLT) were successfully expanded from pediatric to adult cases last 15 years. During this process, graft type has been shifted from left side liver to right side liver. Although the introduction of right lobe graft can successfully increase the actual graft size in LDLT, problem related to "small-for-size grafts" have gradually come to light. "Small-for-size syndrome", such as poor bile production, delayed synthetic function, prolonged cholestasis, and intractable ascites, leading to septic complications and higher mortality, are neither specific nor inevitable in low-weight liver grafts. Many factors other than actual graft weight contribute to the occurrence of "small- for-size syndrome". In the clinical setting, surgical modification targeting portal hemodynamics and tissue congestion is a key to overcome "small-for-size syndrome". Until now, several therapeutic options were reported, but further elucidation of the pathogenesis in "small-for-size syndrome" will be a solution for improving the outcomes in adult-to-adult LDLT.

Humans↗
PubMed2004

Noninvasive evaluation of graft steatosis in living donor liver transplantation.

BACKGROUND: Hepatic steatosis affects graft function as well as postoperative recovery of donors in living donor liver transplantation. Liver macrovesicular steatosis in living donors was assessed using quantitative X-ray computed tomography (CT) analysis and histological examination of intraoperative liver biopsy. METHODS: A total of 266 living donors with complete pretransplant CT data and intraoperative "time 0" biopsy were included in the study. Liver biopsy specimen obtained during donor operation was examined for macrovesicular steatosis and was classified as none; mild (<30%); moderate (30%-60%); or severe (>60%). Liver-to-spleen CT attenuation values ratio (L/S ratio) on noncontrast-CT was evaluated for its usefulness as an index of hepatic steatosis in comparison with other parameters including body mass index (BMI) and serum liver function tests (gamma-glutamyl transpeptidase, alanine aminotransferase, aspartate aminotransferase, cholinesterase, and total cholesterol) using receiver operating characteristic (ROC) analysis. RESULTS.: Histological grade of macrovesicular steatosis was none in 198 patients (74.4%), mild in 50 (18.8%), moderate in 15 (5.7%), and severe in 3 (1.1%). The median L/S ratios for the respective histological grades were 1.20 (range: 1.00-1.46), 1.12 (0.83-1.37), 1.01 (0.74-1.21), and 0.90 (0.70-0.99) (P<0.0001). The ROC curve for L/S ratio was located closest to the upper left corner, and the area under the curve of L/S ratio was significantly larger than that of any other preoperative variables. CONCLUSION: L/S ratio calculated from preoperative CT can be a useful tool to discriminate hepatic macrovesicular steatosis. Based on the present results, the optimal cut-off value for L/S ratio to exclude more than moderate steatosis would be 1.1.

Adult↗
PubMed2004

Ultrastructure of the rostral ventral respiratory group neurons in the ventrolateral medulla of the rat.

The neurons in the ventrolateral medulla that project to the spinal cord are called the rostral ventral respiratory group (rVRG) because they activate spinal respiratory motor neurons. We retrogradely labeled rVRG neurons with Fluoro-Gold (FG) injections into the fourth cervical spinal cord segment to determine their distribution. The rostral half of the rVRG was located in the area ventral to the semicompact formation of the nucleus ambiguus (AmS). A cluster of the neurons moved dorsally and intermingled with the palatopharyngeal motor neurons at the caudal end of the AmS. The caudal half of the rVRG was located in the area including the loose formation of the nucleus ambiguus caudal to the AmS. We also labeled the rVRG neurons retrogradely with wheat germ agglutinin-horseradish peroxidase (WGA-HRP) to determine their ultrastructural characteristics. The neurons of the rVRG were medium to large (38.1 x 22.1 microm), oval or ellipsoid in shape, and had a dark cytoplasm containing numerous free ribosomes, rough endoplasmic reticulum (rER), mitochondria, Golgi apparatuses, lipofuscin granules and a round nucleus with an invaginated nuclear membrane. The average number of axosomatic terminals in a profile was 33.2. The number of axosomatic terminals containing round vesicles and making asymmetric synaptic contacts (Gray's type I) was almost equal to those containing pleomorphic vesicles and making symmetric synaptic contacts (Gray's type II). The axodendritic terminals were large (1.55 microm), and about 60% of them were Gray's type I. The rVRG neurons have ultrastructural characteristics, which are different from the palatopharyngeal motor neurons or the prorpiobulbar neurons.

Animals↗
PubMed2004

Improvement of the MS/MS fragment ion coverage of acidic residue-containing peptides by amidation with 15N-substituted amine.

Tandem mass spectrometry (MS/MS) is a powerful tool for peptide sequencing and characterization. However, the selective cleavage at acidic residues, aspartic acid, and glutamic acid prevents the generation of enough product ions to elucidate the entire sequence. We attempted to solve the problem by converting the residues into the corresponding amides, asparagine and glutamine. The amidation suppressed the cleavage at the converted residues, and the product ions derived from dissociation at other sites became abundant. Incorporation of nitrogen isotope (15)N in the amine constituent for amidation minimized the mass change from -0.984 016 to +0.013 019, allowing easy discrimination of acidic and amide residues in the original sequences by MS/MS database search. In addition, the amidated and unchanged peptides had the same nominal mass, even when the transformation was incomplete, which was approximately 70% in the current condition. The unmodified acidic residues remaining were rather useful to give marker fragments by the dominant dissociation. These results demonstrate that (15)N-amidation is effective in improving the performance of MS/MS to elucidate amino acid sequences of peptides.

Acids↗
PubMed2004

Efficacy of human islet isolation from the tail section of the pancreas for the possibility of living donor islet transplantation.

BACKGROUND: Islet transplantation is on the rise for the treatment of type 1 diabetes. Apparent donor shortages could be alleviated through use of living donor pancreata. A critical issue for using a section of pancreas from living donors is whether islet yields would be sufficient for transplantation. METHODS: After obtaining human pancreata, islets were isolated from the head section (n=20, head group), tail section (n=23, tail group) or whole pancreas (n=24, whole group). Islets were isolated by enzymatic digestion followed by purification, then assessed for yields, purity, morphology, functionality, and insulin content. RESULTS: Fifteen of twenty cases (75%) in the head group, all cases (100%) in the tail group, and 23 of 24 cases (96%) in the whole group were successfully completed for islet isolation. Islet yield per gram pancreas was significantly higher in the tail group compared with both the head and whole groups (head, 1,472+/-326 IE/g; tail, 4,256+/-574 IE/g; whole, 2,424+/-506 IE/g). Total islet yield from the head group was significantly lower compared with both tail and whole groups (head, 75,016+/-18,933 IE; tail, 197,469+/-28,236 IE; whole, 208,207+/-43,414 IE), and the tail group showed similar islet yield to the whole group. The whole group showed significantly lower purities and the head group showed significantly lower morphologic scores. There were no significant differences in viability, function, and insulin content among the three groups. CONCLUSIONS: The tail section of the human pancreas is suitable for islet isolation. The living donor islet transplantation may be feasible using only this section of the pancreas for the first transplantation to reduce hypoglycemic unawareness for small recipients, which might be followed by the second islet transplantation from cadaveric donor.

Adult↗
PubMed2004

Surgical site infection in living-donor liver transplant recipients: a prospective study.

BACKGROUND: Infection is a constant threat for the living-donor liver transplantation (LDLT) recipients, although little information is available on the occurrence of infection in such patients. METHODS: One hundred and thirteen consecutive LDLT recipients were prospectively followed for the presence of surgical site infections (SSIs) defined by CDC from April 2001 to March 2002. Risk factors for SSIs were evaluated by univariate and multivariate analysis. RESULTS: Of the 113 LDLT recipients, 42 (37%) developed 57 episodes of SSIs (21 intraabdominal abscess, 20 peritonitis, 8 cholangitis, and 9 wound). Of the 57 episodes, 29 (51%) had secondary bacteremia in 19 patients. Causative pathogens, including 17 episodes of polymicrobial infections, were 37 gram-positive cocci (17 Staphylococcus aureus, 16 Enterococcus spp., and 4 others), 40 gram-negative rods (25 Enterobacteriaceae, 13 Pseudomonas aeruginosa, and 4 others), and 2 Candida albicans. Univariate analysis revealed that ABO incompatibility and repeat surgery were associated with the development of SSIs. Also, univariate analysis revealed that adult recipients, ABO incompatibility, total operation duration, repeat surgery, and NNIS risk index were associated with secondary bacteremia. Multivariate analysis revealed that ABO incompatibility (OR: 14.0; 95% CI, 2.52-77.2) and repeat surgery (OR: 9.29; 95% CI, 2.00-43.1) were independently associated with secondary bacteremia. Eleven of the 42 cases (26%) who developed SSIs died. Of these 11 cases, 5 (45%) developed secondary bacteremia within 30 days before death. CONCLUSION: SSIs occurred in 37% of LDLT recipients. ABO incompatibility and repeat surgery increased the risk of developing SSIs with secondary bacteremia, which correlated with poor prognosis.

Adolescent↗
PubMed2004

Outcome and risk factors of de novo autoimmune hepatitis in living-donor liver transplantation.

BACKGROUND: Graft dysfunction mimicking autoimmune hepatitis (AIH) develops only rarely after liver transplantation for nonautoimmune liver disease. The long-term prognosis and risk factors of de novo AIH after living-donor liver transplantation (LDLT) are unknown. METHODS: We review our LDLT series to investigate the incidence and outcome of this form of graft dysfunction, focusing on follow-up histology. RESULTS: Of 633 patients who underwent LDLT at Kyoto University from 1990 to 2002, 13 (2.1%) developed graft dysfunction with interface hepatitis resembling AIH (2 males, 11 females). The median age at LDLT of these 13 patients was 10 years (8 months to 26 years). All received tacrolimus-based immunosuppression. The dysfunction presented at a median interval of 3.1 (0.7-9.5) years after LDLT. Nine had definite AIH, and four had probable AIH at the onset of hepatitis. Patients were followed after a median of 3.5 (0.1-8) years from the onset of de novo AIH. Of 11 patients who underwent follow-up histologic evaluation, 3 underwent retransplantation, and 8 continued to have similar findings on subsequent biopsies, with fluctuations in the amount of necroinflammatory activity and an increase in fibrosis despite treatment. In a multivariate analysis, acute rejection episodes and recipient age between 11 and 15 years at LDLT independently had predictive value for the development of de novo AIH. Human leukocyte antigen-A, B, and DR mismatches and sex mismatch did not influence the occurrence of de novo AIH. CONCLUSION: This series highlights the more severe histologic outcome of de novo AIH with longer follow-up despite immunosuppressive treatment. De novo AIH may arise from alloimmunologic injury, marked by clinically obvious episodes of acute rejection.

Adolescent↗
PubMed2004

Induction of antigen-specific immunologic tolerance by in vivo and in vitro antigen-specific expansion of naturally arising Foxp3+CD25+CD4+ regulatory T cells.

Naturally arising CD25(+)CD4(+) regulatory T (T(R)) cells can be exploited to establish immunologic tolerance to non-self antigens. In vivo exposure of CD25(+)CD4(+) T cells from normal naive mice to alloantigen in a T cell-deficient environment elicited spontaneous expansion of alloantigen-specific CD25(+)CD4(+) T(R) cells, which suppressed allograft rejection mediated by subsequently transferred naive T cells, leading to long-term graft tolerance. The expanded T(R) cells, which became CD25(low) in the absence of other T cells, stably sustained suppressive activity, maintained expression levels of other T(R) cell-associated molecules, including Foxp3, CTLA-4 and GITR, and could adoptively transfer tolerance to normal mice. Furthermore, specific removal of the T(R) cells derived from originally transferred CD25(+)CD4(+) T(R) cells evoked graft rejection in the long-term tolerant mice, indicating that any T(R) cells deriving from CD25(-)CD4(+) naive T cells minimally contribute to graft tolerance and that natural T(R) cells are unable to infectiously confer significant suppressive activity to other T cells. Similar antigen-specific expansion of T(R) cells can also be achieved in vitro by stimulating naturally present CD25(+)CD4(+) T cells with alloantigen in the presence of IL-2. The expanded CD25(+)CD4(+) T cells potently suppressed even secondary MLR in vitro and, by in vivo transfer, established antigen-specific long-term graft tolerance. Thus, in vivo or in vitro, direct or indirect ways of antigen-specific expansion of naturally arising Foxp3(+)CD25(+)CD4(+) T(R) cells can establish antigen-specific dominant tolerance to non-self antigens, and would also be instrumental in re-establishing self-tolerance in autoimmune disease and antigen-specific negative control of pathological immune responses.

Adoptive Transfer↗
PubMed2004

The generation of functionally differentiated, three-dimensional hepatic tissue from two-dimensional sheets of progenitor small hepatocytes and nonparenchymal cells.

BACKGROUND: The authors' laboratory has investigated tissue engineering of the liver as a novel approach for treating end-stage liver disease. Fabrication of thick, viable, three-dimensional liver tissue is limited by the lack of vascularity in the tissue-engineered constructs. To overcome this limitation, the authors fabricated three-dimensional, vascularized liver tissue in vivo from two-dimensional cell sheets created from small hepatocytes (SHC) and nonparenchymal cells (NPC) implanted into rat omentum. METHODS: SHC and NPC were cultured on a silicon wafer and lifted as monolayer cell sheets. After maximal hepatotrophic stimulation was induced in the host by injecting retrorsine, creating a portacaval shunt, and performing a partial hepatectomy, these sheets were placed onto the omentum and then rolled into a three-dimensional cylinder. RESULTS: New tissue consisting of both hepatocytes and bile duct-like structures formed by 2 weeks, and the mass of hepatocytes increased in size up to 2 months. The hepatocytes in these constructs were immunohistochemically positive for albumin and transferrin, and bile duct-like structures were positive for gamma-glutamyl transpeptidase, which suggests that they possess liver-specific function. Electron microscopy also revealed structures resembling bile canaliculi. CONCLUSIONS: Functional, morphologically complex new tissue was generated from morphologically simple monolayer cell sheets of SHC and NPC. These results represent an essential step toward the design of tissue-engineered complex vascularized thick tissue.

Animals↗
PubMed2004

Guanidino labeling derivatization strategy for global characterization of peptide mixtures by liquid chromatography matrix-assisted laser desorption/ionization mass spectrometry.

Guanidination performed with isotopic isoforms of O-methylisourea was used in combination with reversed-phase liquid chromatography (LC) matrix-assisted laser desorption/ionization to characterize, both qualitatively and quantitatively, protein mixtures. Synthesis of (13)C- and (15)N(2)-labeled O-methylisourea sulfate produces a molecule that is 3 Da heavier than the light isotopic variant. Protein mixtures containing identical components in different concentration are pooled together following parallel derivatization. Relative quantification of protein mixtures is achieved by mass spectrometry. A difference of 3 Da allows negligible interference between the two isotopic clusters for quantification of peptides up to 1400 Da. Under these conditions, the chromatographic resolution achieved allows separation of different pairs of derivatized peptides without altering the retention time of structurally identical isotopic isoforms. Concomitant isolation of both chemically modified precursors is followed by tandem mass analysis. Activation of the ions via collisions with an inert gas produces isotopically derivatized fragment ions, which appear as doublets in the product ion spectrum. Since the modification occurs on the C-terminal lysine, ions incorporating the guanidino moiety on the C-terminus can be distinguished from those containing the original unmodified peptide N-terminus. Knowledge of the location of the proton can be beneficial to data interpretation and peptide sequencing.

Chromatography, Liquid↗