Objective:To analyze clinical characteristics and genetic characteristics of children with ATP sensitive potassium passage (K ATP-HI). Methods:Forty-five children with genetically confirmed K ATP-HI and their families admitted to Beijing Children′s Hospital of Capital Medical University between February 2002 and December 2018 were selected as the study subjects. A detailed retrospective analysis of the patient's clinical characteristics, diagnosis and treatment process, disease-causing gene carrying status and later follow-up data was performed. ABCC8/KCNJ11 gene was sequenced by second-generation sequencing technology. Results:Among 45 children with K ATP-HI, 34 cases (75.6%) were neonatal onset, the first symptoms of 21 cases (46.7%) were convulsions. 39 cases had been treated with diazoxide, including 12 cases (30.8%) with good efficacy, 16 cases (41%) with poor efficacy and 11 cases with uncertain efficacy. Octreotide was further applied in 18 patients with uncertain or ineffective efficacy after diazoxide treatment, and 13 cases (72.2%) were effective, 3 cases were ineffective, and 2 cases were uncertain. 10 CHI patients who were ineffective to drug treatment or had clearly focal lesions confirmed by 18F-dopa positron emission by computed tomography ( 18F-DOPA PET) scans had undergone surgical treatment, 8 of which underwent partial pancreatectomy and blood glucose returned to normal after the operation; the other 2 cases underwent subtotal pancreatectomy and both had secondary diabetes after operation. Among 45 children with K ATP-HI, 1 case carried both ABCC8 and KCNJ11 mutations, 10 cases carried ABCC8 compound heterozygous mutations, and the remaining 34 cases carried ABCC8/KCNJ11 single genetic mutation. Among them, 21 cases had paternal inheritance, and 3 cases had maternal inheritance, 6 cases were identified with de novo mutations. Conclusions:Diazoxide treatment was ineffective for most K ATP-HI children, but octreotide had a higher effective rate. Partial pancreatectomy for focal type patients had a higher cure rate, and there was a risk of secondary diabetes after subproximal pancreatectomy, so it was very important to clarify the histological type of children before surgery. ABCC8 gene mutations and KCNJ11 gene mutations were the main pathogenic genes of K ATP-HI. Among patients carrying mutations in single ABCC8 or KCNJ11 gene mutation, K ATP-HI inherited by paternity were the majority. Some K ATP-HI children can relieve the hypoglycemia symptoms by themselves.

Objective To analyze the clinical characteristics and gene mutations of 56 patients with congenital hyperinsulinism(CHI)and to provide a theoretical basis for clinical diagnosis and treatment of CHI.Methods Fifty-six children who were diagnosed as CHI between February 2002 and January 2016 in Beijing Children's Hospital Affiliated to Capital Medical University were selected as research subjects.A retrospective study was done about the clinical data and the treatment procedures of the 56 patients,such as perinatal conditions,clinical manifestations,laboratory data,treatments,prognosis and so on.Polymerase chain reaction(PCR)-DNA technology or next-generation sequencing technology was used to analyze the CHI relevant genes of the 56 patients.Results Thirty of the 56 patients carried CHI gene mutation.(1)Twenty-three of 56 patients(41.0％)carried ABCC8/KCNJ11 gene mutations:4 of 23 patients carried complex heterozygous mutation,1 of 23 patients carried both ABCC8 and KCNJ11 gene mutation,1 of 23 patients carried maternally inherited ABCC8 gene mutation,12 of 23 patients carried paternally inherited ABCC8 gene mutation,1 of 23 patients carried paternally inherited KCNJ11 gene mutation,3 of 23 patients carried de novo ABCC8 gene mutation,1 of 23 patients had unknown genetic way,19 of 23 patients were treated with Diazoxide,2 of 19 patients were responsive to Diazoxide,7 of 19 patients were unresponsive to Diazoxide and 10 of 19 patients were uncertain to Diazoxide.(2)Five of 56 patients(8.9％)carried GLUD1 gene mutation,4 of 5 patients were treated with Diazoxide and they were all responsive to Diazoxide.(3)One of 56 patients(1.7％)carried de novo GCK gene mutation,responsive to Diazoxide treatment.(4)One of 56 patients(1.7％)carried maternally inherited SLC16A1 gene mutation,responsive to Diazo-xide treatment.Conclusions The ABCC8 gene and GLUD1 gene mutation are the main causative genes of CHI.The GCK gene and SLC16A1 gene mutation are in the minority.Most ABCC8 gene and KCNJ11 gene mutation are unresponsive to Diazoxide treatment.

Objective To screen the mutation of KATP channel mutations in Chinese pedigrees with infantile onset type 1 diabetes mellitus (T1DM) and neonatal diabetes mellitus.Methods A cohort of 12 children of infant onset T1DM and neonatal diabetes mellitus admitted into Beijing Children's Hospital between March 2004 and June 2013 were selected.PCR amplification and direct sequencing were used to analyze the 39 exons of ABCC8 gene and one exon of KCNJ11.And the mutational sites of the parents of the probands was sequenced in order to identify the inheritance.Results Analysis revealed ABCC8 mutation in 25％ (3/12 cases) of the patients,a case of transient neonatal diabetes (TNDM),a case of permanent neonatal diabetes mellitus (PNDM) and a case of infant onset T1DM.All positive patients showed a known heterozygosis mutation in the ABCC8 gene(R1182Q,c.3545G ＞ A,D209E,c.627C ＞ G,E208K c.622G ＞ A).The residue R1182Q,which was located at a position involved in joining transmembrane domain 2 to nucleotide binding domain 2,the mutations E208K and D209E were located in the intracellular region that links the transmembrane domain with the gatekeeper module.All the three mutations were located throughout the cytoplasm part of SUR1 protein.The TNDM successfully transferred from insulin to oral sulfonylureas therapy.Conclusions There is a complex genetic pathogenesis in neonatal and infant-onset diabetes.The KATP channel activating mutations is one of the main causes of neonatal diabetes mellitus and may cause T1DM in infants in China.Oral Glibenclamide therapy seems highly effective for some patients with the KATP channel activating mutations.

Objective To explore the influence of abnormal donor hepatic artery on hepatic artery and biliary complications after liver transplanta?tion,and summarize the hepatic artery reconstruction procedures during transplantation. Methods The clinical data of 210 cases of liver transplan?tation conducted in our hospital from May 2005 to April 2015 were retrospectively searched for the study,including 42 with abnormal donor hepatic artery. Results Among the 210 liver transplantation,42 cases exhibited abnormal donor hepatic artery,and the aberration rate was 20.0%. Mean volume of blood flow of abnormal group and normal group was 4.7±95.1 mL/min and 190.9±101.6 mL/min,respectively. There was no statistic differ?ence(P=0.519). Twelve cases had arterial complications,the incidence rate was 5.71%,and there was no statistic difference between each group (χ2=0.72,P>0.05). Twenty five cases got biliary complications,the incidence rate was 11.9%,and there was no statistic differences between each group(χ2=0.05,P>0.05). Conclusion There was no statistic difference of mean volume of blood flow after arterial reconstruction between two groups. Liver transplantation with abnormal arterial reconstruction will not increase the incidence rate of arterial and biliary complications.

Objective:To study the effect of immunological molecules expressive state upon the telomerase activation ( TA) of bone marrow mononuclear cells ( BMMNC ) in the hemopoietic microenvironment of patients with immune related hematocytopenia ( IRHS) ,and to explore the immunologic mechanism as well as the clinical significance of hematoclasis in marrow of IRHS patients .Methods:①TRAP-PCR-ELISA method was performed to detect the TA of BMMNC in marrow of 366 IRHS patients before and after therapy.②The molecules HLA-DR,anti-hunman IgG,FcγⅡR,mannose receptor ( MR),IL-17A and its receptor ( IL-17AR) were analysed by immunochemistry and immunofluorescence staining .③The flow cytometric ( FCM) was used to analyse the proportion of CD3+CD4+T cells as well as CD3+CD8+T cells ,CD3-CD19+B cells and CD3-CD16/56+NK cells in peripheral blood lymphocyte.60 cases of health examination were selected as the control group , and 30 cases hypoferric anemia patients were selected as disease control.The differences between patient group and control group were analysed with statistic method .Immunochemistry and immunofluo-rescence staining were performed to in situ analyze the activation-characteristics of immunocyte in bone marrow slides of IRHS ,and the dependablity of cellular immunologic injury was also checked.Results: ①The levels of TA was 0.261 7 ±0.021 6 before treatment , higher than the disease control group (0.061 6±0.031 3 ,P<0.01).Among of them HLA-B27+patients were higher than HLA-B27-patients (0.301 3±0.020 6 vs.0.192 3±0.012 9,P<0.05).Serious IRP patients with HLA-B27+IgG+were obviously higher than HLA-B27-IgG+patients (0.401 6±0.017 2 vs.0.221 1±0.011 0,P<0.01).②In marrow of HLA-B27+IgG+patients,both cell immunity and humoral immunity were in disorder in the hemopoietic microenvironment ,and immonocyte in marrow expressed HLA-DR, FcγⅡR,IL-17A,IL-17RA and MR,and Th,Ts,B cell and NK cell in peripheral blood increased in different degree ,inducing the in-flammation of haemocyte and lead to destruction.③Humoral immunity was in the dominant level in morrow;humoral immunity of HLA-B27-IgG+patients,immonocyte expressed FcγⅡR in high level,but IL-17A was seldom expressed,only CD19+B cell was increased slightly ,the antibody dependent cellular cytotoxicity ( ADCC) was the main mode of destruction.After therapy glucocorticoids associated with ciclosporin A ,the TA level of BMMNC decreased to 0 with devitalization.Conclusion: The telomerase activation of bone marrow mononuclear cells in IRHS is related with the immune state of hemopoietic microenvironment and the pathologic lesion degree of hema -topoietic cell in marrow.It is viral infection and immunological activation as well as a variety of inflammatory factors play a part in the immunologic injury that might be an important factor of the enhancement in TA.

Objective To investigate the biological effect of anti-leukemic cells induced by eosinophilic granulocyte (EOS) in bone marrow of patients with chronic myelogenous leukemia (CML).Methods The BCR-ABL fusion gene as well as the expression of IL-12 and IL-17 mRNA were performed by RT-PCR.The serum concentrations of cytokine IL-12 and IL-17 were determined by enzyme-linked immuno sorbent assay (ELISA).Immunochemistry staining and cytochemistry staining were used to observe the peroxydase (POX) and human Leukocyte antigen (HLA)-DR expression of EOS in bone marrow.Immunofluorescence staining was used to observe mannose receptor (MR),IL-12,IL-17A and IL-17receptor A (IL-17RA) expression of EOS.The results between the CML patients and the healthy controls were compared.Results Serum levels of IL-12 and IL-17 were higher in the 60 CML patients [(196.33 ±21.79) ng/L and (36.55-±3.01) ng/L] than those in the controls [(96.60 ±4.92) ng/L and (23.74 ±1.36) ng/L].In the 32 patients with activated EOS,the levels of IL-12 and IL-17 were (273.12 ± 17.16)ng/L and (40.11 ± 6.13) ng/L,which were significantly higher than those in the non-activated EOS [(126.16 ± 14.27) ng/L and (28.14 ±5.29) ng/L] (P values ＜0.01).IL-12 and IL-17 mRNA were expressed in activated EOS,while BCR-ABL fusion gene was not found.The amounts of EOS were increased abnormally in the bone marrow and peripheral blood of the CML patients with POX positive staining in the cytoplasm and weakly positive HLA-DR staining.It was observed easily by a microscope that EOS could attack leukemic cells in bone marrow through adhesion,capture and phagocytosis.Activated EOS could express IL-12,IL-17A and MR,which was related with the serum levels of these cytokines.Conclusions Activated EOS in bone marrow of CML patients could express IL-12 and IL-17.Activated EOS could induce coup injury to leukemic cell by releasing POX and expressing IL-12 and IL-17.It can also capture or swallow target cells via the expression of MR on the membrane.EOS may play an important role in the anti-tumor immunologic function in bone marrow of CML patients.

Objective Pancreas perfusion is an essential step in human islet isolation.To develop the new methods for introductal canulation,collagenase infusion and to observe their effects on islets isolation.Methods A total of 17 pancreases were digested from March 2005 to April 2010.The pancreases were distended by three methods:the standard method (n =3),the one-cannula method (n =11) and three-cannula method (n =3).In the standard group,the pancreases were completely cut into half at the mid-body.Two catheters were inserted into the main duct:one directed toward the tail and the other to the head.In the one-cannula method group,a long tube was inserted into the duct at the head,advancing to the tail In the three-cannula method group,pancreatic parenchyma was then minimally cut at the mid-body and three catheters were inserted into the main pancreatic duct:one at the head (the first catheter) and two at the mid-body,one toward the tail (the second catheter) and the other toward the head (the third catheter).The pancreases were digested by improved Ricordi technique.Ficoll continuous density/grads centrifuge method was performed to purify the islets.DTZ staining was adopted to identify islets and count islet equivalent (IEQ). AO/EB fluorescence examination was used to count active islet percentage.Static glucose stimulating test (SGS) in vitro was designed to estimate islet function and calculate SI.Results The distension volume of the threecannula method group was 1.24 rnl/g pancreas,and higher than the other groups (for the standard group:0.71 ml/g pancreas; for one-cannula method group:0.96 ml/g pancreas,P＜0.05).The yield of islet in the three-cannula method group and the one-cannula method group was 2514 and 2270 IEQ/g,which was significantly more than that in the standard group (1914 IEQ/g pancreas,P＜0.05).The purity and viability of the islets were 74 ％/79.3 ％,75.6 ％/79.4 ％ and 78.3 ％/84.0 ％ respectively in the three groups with the difference being not significant among the groups.SI in the one cannula method group (4.74) and the three-cannula method group (5.27) was significantly higher than that in the standard group (3.46).ConclusionThe three-cannula method improved collagenase infusion and the islet yields.

Objective To evaluate the effect of islet transplantation for patients with type 2diabetes mellitus (DM). Methods Since December 2007, 4 cases of islet transplantations were performed on 3 patients with type 2 DM and end-stage renal disease (ESRD). Two patients received simultaneous islet-kidney transplant from single-donor (SIK), and one received 2 consecutive islet transplants 5 months following kidney transplantion (IAK). All recipients given insulin with a dose of percutaneous transhepatic portal catheterization. Anti-CD25 monoclonal antibody was used as induction. For SIK, low-doses of Tacrolimus and sirolimus were used as maintenance immunosuppression protocol. For IAK, the maintenance protocol included cyclosporine and MMF.Insulin dose, the level of blood glucose, C-peptide and the value of HbA1 were observed. Results The first patient of SIKhad normal glucose level 3 days after surgery and became insulin independent within the first month, but insulin was administered gradually and the dose reduced to 1/3. The second patient of SIK died of bleeding and secondary infection of liver puncture site 5 days following operation, the blood glucose level recovered to normal 24 h after operation. The insulin dose of the patient of IAK was reduced to 1/2 after the first transplant. The patient became insulin free after the second operation. The level of fasting and postprandial C-peptide of the surviving recipients increased by 600 pmol/L. The value of HbA1 of the SIK was 6.7 %～7.3 %, while that of the IAK was 5. 5 %～ 5. 9 %. Conclusion Islet transplantation is an effective treatment for patients with type 2 DM.

Objective To summarize our experience in the liver transplantation from the donation after cardiac death (DCD).Methods The livers from three DCD donors (2 cases of brain trauma and 1 case of cerebral hemorrhage) were harvested according to the Guidelines for Donation after Cardiac Death in China.These grafts were orthotopically transplanted into three recipients including 2 cases of decompensative hepatic cirrhosis and 1 case of primary liver cancer.The warm ischemic time ranged from 7.5 to 10 min and the cold ischemic time was 4.5,8.2 and 6.5 h respectively.Postoperative immunosuppressive regimens included prednisone,FK506 and mycophenolate mofetil (MMF).Antibiotics and anticoagulatants were used accordingly.Results All of the 3 recipients obtained normal liver function within 3 weeks since the grafts were implanted without PNF,thrombosis and rejection.No postoperative complications occurred in 3 recipients during the follow-up period of 2 to 9 months with normal liver function.Conclusion The liver transplant from DCD donor showed good results in our center.Chinese group Ⅲ of DCD donor,UW score above the middle level and the short warm ischemic time are three keys ensuring the success of the liver transplant from DCD donors.

Objective To summarize the clinical experience of harvesting and using the kidneys from donation after cardiac death (DCD) donors.Methods Fourteen kidney transplantations were successfully performed on 14 patients with end-stage renal diseases.The kidneys were harvested from 7 volunteer donors (age 30～53 years) diagnosed with cardiac death,who were scored 19～23according to the University of Wisconsin donation after cardiac death evaluation.Primary diseases of the donors were cerebral hemorrhage,brain injury,ischemic cerebral vascular disease and brain tumor.Warm ischemia time ranged from 5 to 45 min,and cold ischemia time was 4.5 ～ 12.5 h.Results After transplantation,three patients had delayed graft function (DGF),one had primary non-function (PNF),and two patients developed acute rejection.In the patient with PNF,the transplanted kidney was removed one day after operation and the patient went back to hemodialysis.One patient with DGF was still in recovery with serum creatine 149 μmnol/L (within 3 months after operation).The above two cases both utilized the kidneys with 45 min of warm ischemia time.The rest 12 patients were discharged with normal renal function.Conclusion Under the condition of our country,kidneys strictly harvested from DCD donors can be used as one of the main sources of kidney grafts for kidney transplantation.