AIM: Artificial beta-cell lines may offer an abundant source of cells for the treatment of type I diabetes, but insulin secretion in beta-cells is tightly regulated in physiological conditions. The Tet-On system is a ...AIM: Artificial beta-cell lines may offer an abundant source of cells for the treatment of type I diabetes, but insulin secretion in beta-cells is tightly regulated in physiological conditions. The Tet-On system is a "gene switch" system, which can induce gene expression by administration of tetracycline (Tet) derivatives such as doxcycline (Dox). Using this system, we established 293 cells to an artificial cell line secreting insulin in response to stimulation by Dox. METHODS: The mutated proinsulin cDNA was obtained from plasmid pcDNA3.1/C-mINS by the polymerase chain reaction (PCR), and was inserted downstream from the promoter on the expression vector pTRE2, to construct a recombined expression vector pTRE2mINS. The promoter on pTRE2 consists of the tetracycline-response element and the CMV minimal promoter and is thus activated by the reverse tetracycline-controlled transactivator (rtTA) when Dox is administrated. pTRE2mINS and plasmid pTK-Hyg encoding hygromycin were co-transfected in the tet293 cells, which express rtTA stably. Following hygromycin screening, the survived cells expressing insulin were selected and enriched. Dox was used to control the expression of insulin in these cells. At the levels of mRNA and protein, the regulating effect of Dox in culture medium on the expression of proinsulin gene was estimated respectively with Northern blot, RT-PCR, and radioimmunoassay. RESULTS: From the 28 hygromycin-resistant cell strains, we selected one cell strain (tet293/Ins6) secreting insulin not only automatically, but in response to stimulation by Dox. The amount on insulin secretion was dependent on the Dox dose (0,10,100,200,400,800 and 1000 microg.L(-1)), the level of insulin secreted by the cells treated with Dox (1000 microg.L(-1)) was 241.0pU.d(-1).cell(-1) , which was 25-fold that of 9.7pU.d(-1).cell(-1) without Dox treatment. Northern blot analyses and RT-PCR further confirmed that the transcription of insulin gene had already been up-regulated after exposing tet293/Ins6 cells to Dox 展开更多
OBJECTIVE: To review the current progress of islet cell transplantation in patients with insulin-dependent diabetes, emphasizing on the difficulties with recovering and preserving islet cell mass and function, 30% of ...OBJECTIVE: To review the current progress of islet cell transplantation in patients with insulin-dependent diabetes, emphasizing on the difficulties with recovering and preserving islet cell mass and function, 30% of which is lost during the peri-transplantation period. RESULTS: The islet-cell isolation technique is perfected, but improvements are still progressing in two major directions: preservation of islet cells and tolerance induction. Optimum islet cell viability and function depends on appropriate revascularization of the islet graft and blockade of thrombus formation as well as cytokine and free radical release. Conditioning the islet cells in-vitro prior to transplantation to either upregulate VEGF expression or downregulate NF-kappa B transcription factor has proven to improve revascularization and to prevent islet cell apoptosis and cytokine-mediated damage. Tolerance induction is currently being best achieved by selecting and combining immunosuppressive agents such as monoclonal antibodies which target the major signaling molecules during immune activation, but which are least toxic to islet cells. CONCLUSIONS: Patients with insulin-dependent diabetes will greatly benefit from current developments in effective approaches to protect islets during the peritransplant period. Emerging interest in stem cell biology and differentiation may provide the ultimate solution to the problem of organ scarcity and islet cell protection from the peritransplant induced damage.展开更多
Bone marrow mesenchymal stem cells (BMSCs) have the ability of self-renewaland multi-directional differentiation. Recent reports showed that BMSCs could differentiate intoendocrine cells of pancreas. However, the diff...Bone marrow mesenchymal stem cells (BMSCs) have the ability of self-renewaland multi-directional differentiation. Recent reports showed that BMSCs could differentiate intoendocrine cells of pancreas. However, the differentiation is not efficient enough to produceinsulin-producing cells for the future therapeutic use. Pdx-1 is a crucial regulator for pancreaticdevelopment. Therefore we constructed a eukaryotic expression vector containing Pdx-1 to determinethe effect of Pdx-1 expression on differentiation of BMSCs in vitro. The results showed that BMSCscould self-assemble to form functional pancreatic islet-like structures after differentiation invitro. The proportion of insulin-producing cells differentiated from Pdx-1 +BMSCs was 28.23%+-2.56%,higher than that from BMSCs transfected with vacant vector and Pdx-1'' BMSCs (7.23%+-1.56% and4.08%+-2.69% respectively) by flow cytometry. Immunocytochemical examination also testified theexpression of multiple bate-cells-specific genes such as insulin, glucagons, somatostatin indifferentiated BMSCs. The results also revealed that the expressions of genes mentioned above inPdx-1+BMSCs were higher than that in Pdx-VBMSCs, which was confirmed by Western blotting analysisand RT-PCR. Glucose-inducedinsulin secretion from Pdx-1+BMSCs in 5mmol/L and 25mmol/L glocuse was(56.61 +-4.82) uU/ml and (115.29+-2.56) uU/ml respectively, which were much higher than those fromPdx-1 BMSCs((25.53 +-6.49) uU/mL and (53.26 + 7.56) uU/mL respectively). Grafted animals were ableto maintain their body weight and survive for relatively longer periods of time than hyperglycemicsham-grafted controls, which demonstrated an overall beneficial effect of the grafted cells on thehealth of the animals. These findings thus suggested that exogenous expression of Pdx-1 shouldprovide a promising approach for efficiently producing islet-like cells from BMSCs for the futuretherapeutic use in diabetic patients.展开更多
Background Islet transplantation represents an ideal therapeutic approach for treatment of type 1 diabetes but islet function and regeneration may be influenced by necrosis or apoptosis induced by oxidative stress and...Background Islet transplantation represents an ideal therapeutic approach for treatment of type 1 diabetes but islet function and regeneration may be influenced by necrosis or apoptosis induced by oxidative stress and other insults. Heme oxygenase-1 （HO-1） is the rate-limiting enzyme in the catabolism of heme into biliverdin, releasing free iron and carbon monoxide. It has also been reported to be an antioxidant enzyme which can improve the function of grafted islets by cytoprotection via free radical scavenging and apoptosis prevention. In the present study, we investigated whether transduction of HO-1 genes into human islets with an adenovirus vector has cytoprotective action on islets cultured in vitro and discuss this method of gene therapy for clinical islet transplantation. Methods Cadaveric pancreatic islets were isolated and purified in vitro. Transduction efficiency of islets was determined by infecting islets with adenovirus vector containing the enhanced green fluorescent protein gene （Ad-EGFP） at multiplicities of infection （MOI） of 2, 5, 10, or 20. Newly isolated islets were divided into three groups： EGFP group, islets transduced with Ad-EGFP using MOI=20; HO-1 group, transduced with adenovirus vectors containing the human HO-1 gene using MOI=20; and control group, mock transduced islets. Insulin release after glucose stimulation of the cell lines was determined by a radioimmunoassay kit and the stimulation index was calculated. Flow cytometry was used to detect apoptotic cells in the HO-1 group and in the control group after induction by recombinant human tumor necrosis factor-α （rTNFα） and cycloheximide （CHX） for 48 hours. Results Adenovirus vectors have a high efficiency of gene transduction into adult islet cells. Transduction of islets with the Ad-EGFP was most successful at MOI 20, at which MOI fluorescence was very intense on day 7 after transduction and EGFP was expressed in cultured islet cells for more than four weeks in vitro. The insulin release in the control gro展开更多
AIM： To investigate the ability of a genetically altered embryonic stem （ES） cell line to generate insulin-producing cells in vitro following transfer of the Nkx2.2 gene.METHODS： Hamster Nkx2.2 genes were transfer...AIM： To investigate the ability of a genetically altered embryonic stem （ES） cell line to generate insulin-producing cells in vitro following transfer of the Nkx2.2 gene.METHODS： Hamster Nkx2.2 genes were transferred into mouse ES cells. Parental and Nkx2.2-transfected ES cells were initiated toward differentiation in embryoid body （EB） culture for 5 d and the resulting EBs were transferred to an attached culture system. Dithizone （DTZ）, a zincchelating agent known to selectively stain pancreatic beta cells, was used to detect insulin-producing cells.The outgrowths were incubated in DTZ solution （final concentration, 100μg/mL） for 15 rain before being examined microscopically. Gene expression of the endocrine pancreatic markers was also analyzed by RT-PCR. In addition, insulin production was determined immunohistochemically and its secretion was examined using an ELISA.RESULTS： DTZ-stained cellular clusters appeared after approximately 14 d in the culture of Nkx2.2-transfected ES cells （Nkx-ES cells）, which was as much as 2 wk earlier, than those in the culture of parental ES cells （wt-ES）. The frequency of DTZ-positive cells among total cultured cells on day 28 accounted for approximately 1.0% and 0.1% of the Nkx-ES- and wt-ES-derived EB outgrowths, respectively. The DTZ-positive cellular clusters were found to be immunoreactive to insulin, while the gene expressions of pancreatic-duodenal homeobox 1 （PDX1）, proinsulin 1 and proinsulin 2 were observed in the cultures that contained DTZ-positive cellular clusters.Insulin secretion was also confirmed by ELISA, whereas glucose-dependent secretion was not demonstrated.CONCLUSION： Nkx2.2-transfected ES cells showed an ability to differentiate into insulin-producing cells.展开更多
Background Costimulatory signals play a vital role in T cell activation. Blockade of costimulatory pathway by CTLA4Ig or CD40LIg have enhanced graft survival in experimental transplantation models yet mechanisms remai...Background Costimulatory signals play a vital role in T cell activation. Blockade of costimulatory pathway by CTLA4Ig or CD40LIg have enhanced graft survival in experimental transplantation models yet mechanisms remain undetermined.We investigated the effects of CTLA4Ig and CD40LIg gene transfer on islet xenografts rejection in rats.Methods Human islets were infected with recombinant adenoviruses containing CTLA4Ig and CD40LIg genes and implanted beneath the kidney capsule of diabetic rats. Levels of blood sugar, morphological changes, and survival of grafts were recorded. Expressions of CTLA4Ig, CD40LIg and insulin were detected by immunohistochemical staining and cytokines levels were quantified by enzyme-linked immunosorbent assay （ELISA）.Results Blood glucose levels in transplant rats decreased to normal level on the 2nd day post transplantation. The mean blood glucose in the control group, CTLA4Ig transfected group, CD40LIg transfected group and CTLA4Ig ＋CD40LIg cotransfected group increased on days 8, 24, 21, 68, post transplantation respectively. The grafts in control group, CTLA4Ig transfected group, CD40LIg transfected group and CTLA4Ig ＋ CD40LIg cotransfected group survived for （8±1）, （29±4）, （27±3）, and （74±10） days, respectively. Survival in CTLA4Ig ＋ CD40LIg cotransfected group was significantly longer. Survivals of CTLA4Ig transfected group and CD40LIg transfected group were significantly longer than control group. In controJ animals, serum interleukin-2 and tumor necrosis factor a concentration significantly increased within seven days post transplantation. Haematoxylin eosin staining of grafts showed live islets in situ of transplant rats without inflammatory cell infiltration. Immunohistochemical staining confirmed the expression of insulin at islets in all experimental groups.Conclusions Transfer of CTLA4Ig and CD40Llg genes, especially the cotransfer of both, inhibits rejection of murine islet xenografts. Downregulated expressions of Th1 cells related cytokines mig展开更多
At present, proven clinical treatments but no cures are available for diabetes, a global epidemic with a huge economic burden. Transplantation of islets ofLangerhans by their infusion into vascularized organs is an ex...At present, proven clinical treatments but no cures are available for diabetes, a global epidemic with a huge economic burden. Transplantation of islets ofLangerhans by their infusion into vascularized organs is an experimental clinical protocol, the first approach to attain cure. However, it is associated with lifelong use of immunosuppressants. To overcome the need for immunosuppression, islets are encapsulated and separated from the host immune system by a permselective membrane. The lead material for this application is alginate which was tested in many animal models and a few clinical trials. This review discusses all aspects related to the function of transplanted encapsulated islets such as the basic requirements from a permselective membrane(e.g., allowable hydrodynamic radii, implications of the thickness of the membrane and relative electrical charge). Another aspect involves adequate oxygen supply, which is essential for survival/performance of transplanted islets, especially when using large retrievable macrocapsules implanted in poorly oxygenated sites like the subcutis. Notably, islets can survive under low oxygen tension and are physiologically active at > 40 Torr. Surprisingly, when densely crowded, islets are fully functional under hyperoxic pressure of up to 500 Torr(> 300% of atmospheric oxygen tension). The review also addresses an additional category of requirements for optimal performance of transplanted islets, named auxiliary technologies. These include control of inflammation, apoptosis, angiogenesis, and the intra-capsular environment. The review highlights that curing diabetes with a functional bio-artificial pancreas requires optimizing all of these aspects, and that significant advances have already been made in many of them.展开更多
Erdheim-Chester Disease (ECD) is a rare condition and has various differential diagnoses with other forms of histiocytosis, classified as one of non-Langerhans histiocytosis. The diagnosis of this condition remains ch...Erdheim-Chester Disease (ECD) is a rare condition and has various differential diagnoses with other forms of histiocytosis, classified as one of non-Langerhans histiocytosis. The diagnosis of this condition remains challenging because its presentation includes non-specific systemic manifestations that can affect different organs caused by deposition of lipids and fibrosis. Most common include bone pain followed by progressive weakness and different lung manifestations. This case is about a rare presentation of ECD with Langerhans Histiocytosis as overlap syndrome, with findings of both diseases in a middle aged woman that presented dyspnea as the first symptom. The patient was treated initially as heart failure and remained without any improvement, being admitted to investigate. After a stricted follow-up, bone and lung involvement were noticed and a skin biopsy unveiled xanthomatized macrophages accompanied by Touton giant cells. This condition remains an important clinical entity and should provide new insights for clinicians dealing with respiratory diseases.展开更多
BACKGROUND Langerhans cell histiocytosis(LCH)is characterized by diabetes insipidus and is an uncommon occurrence.Pathological biopsies still have a certain degree of diagnostic probability.We present a case in which ...BACKGROUND Langerhans cell histiocytosis(LCH)is characterized by diabetes insipidus and is an uncommon occurrence.Pathological biopsies still have a certain degree of diagnostic probability.We present a case in which LCH initially affected the pituitary gland.This resulted in a misdiagnosis of chronic inflammation upon pathological examination.CASE SUMMARY A 25-year-old female exhibited symptoms of diabetes insipidus.Magnetic resonance imaging revealed an enhanced foci in the pituitary gland.After surgical resection of the pituitary lesion,the pathological diagnosis was chronic inflam-mation.However,the patient later experienced bone destruction in the skull and lower limb bones.After the lower limb bone lesion was compared with the initial pituitary lesion,the final diagnosis was modified to LCH.The patient was treated with multiple chemotherapy courses.However,the patient’s condition gradually worsened,and she eventually passed away at home.CONCLUSION LCH should be considered when patients exhibit diabetes insipidus and absence of high signal intensity in the pituitary gland on sagittal T1-weighted image and abnormal enhancement in the pituitary region.展开更多
BACKGROUND Langerhans cell histiocytosis(LCH)is a histiocytic proliferative disease caused by clonal proliferation of Langerhans cells,which is currently defined as an inflam-matory myeloid tumor.It is rare in adults,...BACKGROUND Langerhans cell histiocytosis(LCH)is a histiocytic proliferative disease caused by clonal proliferation of Langerhans cells,which is currently defined as an inflam-matory myeloid tumor.It is rare in adults,with an incidence of 1–2 per million,and is highly heterogeneous in clinical presentation,with unpredictable disease progression and outcome.CASE SUMMARY A 52-year-old postmenopausal female patient presented to the gynecology department in July 2023 with bilateral vulvar masses.She was diagnosed with recurrent multisystem LCH.The patient had previously been diagnosed with a single-system and single-focal LCH in October 2021 due to a right maxillofacial mass,which resolved after surgical treatment.A chemotherapy regimen was developed after multidisciplinary consultation.Six cycles of chemotherapy resulted in partial remission,and maintenance chemotherapy is currently being administered.CONCLUSION Recurrent LCH involving the bilateral vulva has been poorly reported.Compre-hensive imaging and pathological evaluation is important for diagnosis.The model of joint multidisciplinary specialist diagnosis and treatment is worthy of clinical application.展开更多
Background The sensitization and elicitation phases are involved in the immunopathogenesis of contact hypersensitivity （CHS）. Langerhans cells （LCs） are believed to play pivotal roles in the sensitization stage of...Background The sensitization and elicitation phases are involved in the immunopathogenesis of contact hypersensitivity （CHS）. Langerhans cells （LCs） are believed to play pivotal roles in the sensitization stage of CHS. Local hyperthermia on skin induces the migration as well as maturation of epidermal LCs. Although fever-range whole body hyperthermia and local hyperthermia at 43℃ prior to sensitization were reported to suppress CHS, the effects of different temperatures and the timing sequence of local hyperthermia on CHS have not been tackled. Methods Local hyperthermia was applied to murine dorsal skin 3 days prior to, concurrent with, or 2 days post sensitization with fluorescein isothiocyanate （FITC） in BALB/c mice. Local hyperthermia temperatures at 37℃, 39℃, 41℃ and 43℃ were applied to mouse dorsal skin and the severity of CHS was calculated by measuring the swelling response of the challenged ears. Results Local hyperthermia at 39℃, 41℃ and 43℃ prior to sensitization reduced the severity of CHS, as compared with that at 37℃. The suppression of CHS was temperature dependant in that higher temperature had a stronger effect. On the contrary, the hyperthermia treatments, either concurrent with or post-sensitization, resulted in an enhanced temperature-dependant ear swelling response. Conclusions The severity of murine CHS could be influenced by local hyperthermia at the sensitization stage in a temperature dependant manner. The temporal effect of local hyperthermia suggested a novel factor in interpreting the severity of allergic contact dermatitis.展开更多
Langerhans cell histiocytosis(LCH) is a rare syndrome characterized by unifocal,multifocal unisystem,or disseminated/multi-system disease that commonly involves the bone,skin,lymph nodes,pituitary,or sometimes lung(al...Langerhans cell histiocytosis(LCH) is a rare syndrome characterized by unifocal,multifocal unisystem,or disseminated/multi-system disease that commonly involves the bone,skin,lymph nodes,pituitary,or sometimes lung(almost exclusively in smokers) causing a variety of symptoms from rashes and bone lesions to diabetes insipidus or pulmonary infiltrates.We present a previously unreported case of gastrointestinal LCH as well as a novel characteristic lesion affecting the colon of a young woman who presented with signs and symptoms mimicking acute on chronic appendicitis.Immunohistochemical analysis of appendectomy specimen and nodular specimens on colonoscopy demonstrated S-100,CD1a,and langerin reactivity.The patient underwent systemic chemotherapy with cytarabine and demonstrated excellent response to therapy.展开更多
Background： Pulmonary Langerhans cell histiocytosis （PLCH） is an interstitial primary pulmonary disease, characterized by Langerhans cell proliferation. It is easily misdiagnosed in children. This study aimed to ch...Background： Pulmonary Langerhans cell histiocytosis （PLCH） is an interstitial primary pulmonary disease, characterized by Langerhans cell proliferation. It is easily misdiagnosed in children. This study aimed to characterize the clinical manifestations and features of PLCH by retrospective analysis. Meihods： A retrospective analysis was performed in 117 PLCH patients out of 338 LCH patients who were admitted in our center from November 2006 to October 2013. Variables between two groups were compared by Mann-Whitney U-test and Chi-square test. Kaplan-Meier curves were constructed to compare the survival rates and Cox regression to evaluate the effect of risk factors. Results： The median age of PLCH group was significantly lower than that ofnon-PkCH group （ 18.63 months vs. 43.4 months, P 〈 0.001 ）. All PLCH children had other organ involvement and only 11 cases （9.4%） had respiratory symptoms. The most common radiologic finding was cystic lesions （29 cases, 24.8%）. Pulmonary thnction abnormalities were dominated by obstructive ventilatory dysfunction （63 cases, 82.9%）. The 5-year overall survival （OS） of PECH children was 93.6% ± 2.3% and the event-free survival （EFS） was 55.7% ± 5.2%. Among the 38 cases with progressed or relapsed disease, five cases （13.2%） were due to progression or recurrence of lung damage. The 5-year OS of PECH children with ＂risk organ＂ involvement was significantly lower than those without ＂risk organ＂ involvement （86.0% ± 4.9% vs. 100%,X^2 = 8.793, P= 0.003）. The difference of EFS between two groups was also significant （43.7% ± 7.7% vs. 66.3% ± 6.5%,X^2 = 5.399, P = 0.020）. The ＂＇risk organ＂ involvement had a significant impact on survival （hazard ratio = 1.9, P = 0.039）. Conclusions： PLCH mainly occurs in young children, and only a small percentage of patients have respiratory symptoms. They generally have other organ involvement. Most of PLCH children have a good prognosis and most lung lesions could have improved展开更多
Pulmonary Langerhans＇ cell histiocytosis （PLCH） is an isolated form of Langerhans＇ cell histiocytosis thatusually develops in cigarette smokers.1＇2^ PLCH usually has typical light microscopic morphology with a sp...Pulmonary Langerhans＇ cell histiocytosis （PLCH） is an isolated form of Langerhans＇ cell histiocytosis thatusually develops in cigarette smokers.1＇2^ PLCH usually has typical light microscopic morphology with a spectrum of progressive changes.2-4^ It is characterized by peribronchiolar Langerhans＇ cell infiltrates mixed with macrophages, lymphocytes and eosinophils, that form stellate nodules. The nodular lesions are frequently cavitated and form thick- and thin-walled cysts, which are thought to represent enlarged airway lumina. PLCH lesions display temporal microscopic heterogeneity, with a progression from dense cellular nodules to cavitary nodules with increasing degrees of fibrosis that may extend along alveolar walls. In advanced cases,展开更多
基金the"Hundred Talents"Program of Shanghai Municipal Government,No.98BR018
文摘AIM: Artificial beta-cell lines may offer an abundant source of cells for the treatment of type I diabetes, but insulin secretion in beta-cells is tightly regulated in physiological conditions. The Tet-On system is a "gene switch" system, which can induce gene expression by administration of tetracycline (Tet) derivatives such as doxcycline (Dox). Using this system, we established 293 cells to an artificial cell line secreting insulin in response to stimulation by Dox. METHODS: The mutated proinsulin cDNA was obtained from plasmid pcDNA3.1/C-mINS by the polymerase chain reaction (PCR), and was inserted downstream from the promoter on the expression vector pTRE2, to construct a recombined expression vector pTRE2mINS. The promoter on pTRE2 consists of the tetracycline-response element and the CMV minimal promoter and is thus activated by the reverse tetracycline-controlled transactivator (rtTA) when Dox is administrated. pTRE2mINS and plasmid pTK-Hyg encoding hygromycin were co-transfected in the tet293 cells, which express rtTA stably. Following hygromycin screening, the survived cells expressing insulin were selected and enriched. Dox was used to control the expression of insulin in these cells. At the levels of mRNA and protein, the regulating effect of Dox in culture medium on the expression of proinsulin gene was estimated respectively with Northern blot, RT-PCR, and radioimmunoassay. RESULTS: From the 28 hygromycin-resistant cell strains, we selected one cell strain (tet293/Ins6) secreting insulin not only automatically, but in response to stimulation by Dox. The amount on insulin secretion was dependent on the Dox dose (0,10,100,200,400,800 and 1000 microg.L(-1)), the level of insulin secreted by the cells treated with Dox (1000 microg.L(-1)) was 241.0pU.d(-1).cell(-1) , which was 25-fold that of 9.7pU.d(-1).cell(-1) without Dox treatment. Northern blot analyses and RT-PCR further confirmed that the transcription of insulin gene had already been up-regulated after exposing tet293/Ins6 cells to Dox
文摘OBJECTIVE: To review the current progress of islet cell transplantation in patients with insulin-dependent diabetes, emphasizing on the difficulties with recovering and preserving islet cell mass and function, 30% of which is lost during the peri-transplantation period. RESULTS: The islet-cell isolation technique is perfected, but improvements are still progressing in two major directions: preservation of islet cells and tolerance induction. Optimum islet cell viability and function depends on appropriate revascularization of the islet graft and blockade of thrombus formation as well as cytokine and free radical release. Conditioning the islet cells in-vitro prior to transplantation to either upregulate VEGF expression or downregulate NF-kappa B transcription factor has proven to improve revascularization and to prevent islet cell apoptosis and cytokine-mediated damage. Tolerance induction is currently being best achieved by selecting and combining immunosuppressive agents such as monoclonal antibodies which target the major signaling molecules during immune activation, but which are least toxic to islet cells. CONCLUSIONS: Patients with insulin-dependent diabetes will greatly benefit from current developments in effective approaches to protect islets during the peritransplant period. Emerging interest in stem cell biology and differentiation may provide the ultimate solution to the problem of organ scarcity and islet cell protection from the peritransplant induced damage.
基金financially supported by the National Natural Science Foundation of China(Grant No.30200128).
文摘Bone marrow mesenchymal stem cells (BMSCs) have the ability of self-renewaland multi-directional differentiation. Recent reports showed that BMSCs could differentiate intoendocrine cells of pancreas. However, the differentiation is not efficient enough to produceinsulin-producing cells for the future therapeutic use. Pdx-1 is a crucial regulator for pancreaticdevelopment. Therefore we constructed a eukaryotic expression vector containing Pdx-1 to determinethe effect of Pdx-1 expression on differentiation of BMSCs in vitro. The results showed that BMSCscould self-assemble to form functional pancreatic islet-like structures after differentiation invitro. The proportion of insulin-producing cells differentiated from Pdx-1 +BMSCs was 28.23%+-2.56%,higher than that from BMSCs transfected with vacant vector and Pdx-1'' BMSCs (7.23%+-1.56% and4.08%+-2.69% respectively) by flow cytometry. Immunocytochemical examination also testified theexpression of multiple bate-cells-specific genes such as insulin, glucagons, somatostatin indifferentiated BMSCs. The results also revealed that the expressions of genes mentioned above inPdx-1+BMSCs were higher than that in Pdx-VBMSCs, which was confirmed by Western blotting analysisand RT-PCR. Glucose-inducedinsulin secretion from Pdx-1+BMSCs in 5mmol/L and 25mmol/L glocuse was(56.61 +-4.82) uU/ml and (115.29+-2.56) uU/ml respectively, which were much higher than those fromPdx-1 BMSCs((25.53 +-6.49) uU/mL and (53.26 + 7.56) uU/mL respectively). Grafted animals were ableto maintain their body weight and survive for relatively longer periods of time than hyperglycemicsham-grafted controls, which demonstrated an overall beneficial effect of the grafted cells on thehealth of the animals. These findings thus suggested that exogenous expression of Pdx-1 shouldprovide a promising approach for efficiently producing islet-like cells from BMSCs for the futuretherapeutic use in diabetic patients.
基金This work was supported by grants from the National Natural Science Foundation of China (No. 30571759), Social Development Foundation of Shanghai, Important Item Foundation of Fujian Scientific and Technical Committee (No. 2002Y007), and Important Item Foundation of "Tenth Five Program" of the Army (No. 04Z007).
文摘Background Islet transplantation represents an ideal therapeutic approach for treatment of type 1 diabetes but islet function and regeneration may be influenced by necrosis or apoptosis induced by oxidative stress and other insults. Heme oxygenase-1 （HO-1） is the rate-limiting enzyme in the catabolism of heme into biliverdin, releasing free iron and carbon monoxide. It has also been reported to be an antioxidant enzyme which can improve the function of grafted islets by cytoprotection via free radical scavenging and apoptosis prevention. In the present study, we investigated whether transduction of HO-1 genes into human islets with an adenovirus vector has cytoprotective action on islets cultured in vitro and discuss this method of gene therapy for clinical islet transplantation. Methods Cadaveric pancreatic islets were isolated and purified in vitro. Transduction efficiency of islets was determined by infecting islets with adenovirus vector containing the enhanced green fluorescent protein gene （Ad-EGFP） at multiplicities of infection （MOI） of 2, 5, 10, or 20. Newly isolated islets were divided into three groups： EGFP group, islets transduced with Ad-EGFP using MOI=20; HO-1 group, transduced with adenovirus vectors containing the human HO-1 gene using MOI=20; and control group, mock transduced islets. Insulin release after glucose stimulation of the cell lines was determined by a radioimmunoassay kit and the stimulation index was calculated. Flow cytometry was used to detect apoptotic cells in the HO-1 group and in the control group after induction by recombinant human tumor necrosis factor-α （rTNFα） and cycloheximide （CHX） for 48 hours. Results Adenovirus vectors have a high efficiency of gene transduction into adult islet cells. Transduction of islets with the Ad-EGFP was most successful at MOI 20, at which MOI fluorescence was very intense on day 7 after transduction and EGFP was expressed in cultured islet cells for more than four weeks in vitro. The insulin release in the control gro
文摘AIM： To investigate the ability of a genetically altered embryonic stem （ES） cell line to generate insulin-producing cells in vitro following transfer of the Nkx2.2 gene.METHODS： Hamster Nkx2.2 genes were transferred into mouse ES cells. Parental and Nkx2.2-transfected ES cells were initiated toward differentiation in embryoid body （EB） culture for 5 d and the resulting EBs were transferred to an attached culture system. Dithizone （DTZ）, a zincchelating agent known to selectively stain pancreatic beta cells, was used to detect insulin-producing cells.The outgrowths were incubated in DTZ solution （final concentration, 100μg/mL） for 15 rain before being examined microscopically. Gene expression of the endocrine pancreatic markers was also analyzed by RT-PCR. In addition, insulin production was determined immunohistochemically and its secretion was examined using an ELISA.RESULTS： DTZ-stained cellular clusters appeared after approximately 14 d in the culture of Nkx2.2-transfected ES cells （Nkx-ES cells）, which was as much as 2 wk earlier, than those in the culture of parental ES cells （wt-ES）. The frequency of DTZ-positive cells among total cultured cells on day 28 accounted for approximately 1.0% and 0.1% of the Nkx-ES- and wt-ES-derived EB outgrowths, respectively. The DTZ-positive cellular clusters were found to be immunoreactive to insulin, while the gene expressions of pancreatic-duodenal homeobox 1 （PDX1）, proinsulin 1 and proinsulin 2 were observed in the cultures that contained DTZ-positive cellular clusters.Insulin secretion was also confirmed by ELISA, whereas glucose-dependent secretion was not demonstrated.CONCLUSION： Nkx2.2-transfected ES cells showed an ability to differentiate into insulin-producing cells.
文摘Background Costimulatory signals play a vital role in T cell activation. Blockade of costimulatory pathway by CTLA4Ig or CD40LIg have enhanced graft survival in experimental transplantation models yet mechanisms remain undetermined.We investigated the effects of CTLA4Ig and CD40LIg gene transfer on islet xenografts rejection in rats.Methods Human islets were infected with recombinant adenoviruses containing CTLA4Ig and CD40LIg genes and implanted beneath the kidney capsule of diabetic rats. Levels of blood sugar, morphological changes, and survival of grafts were recorded. Expressions of CTLA4Ig, CD40LIg and insulin were detected by immunohistochemical staining and cytokines levels were quantified by enzyme-linked immunosorbent assay （ELISA）.Results Blood glucose levels in transplant rats decreased to normal level on the 2nd day post transplantation. The mean blood glucose in the control group, CTLA4Ig transfected group, CD40LIg transfected group and CTLA4Ig ＋CD40LIg cotransfected group increased on days 8, 24, 21, 68, post transplantation respectively. The grafts in control group, CTLA4Ig transfected group, CD40LIg transfected group and CTLA4Ig ＋ CD40LIg cotransfected group survived for （8±1）, （29±4）, （27±3）, and （74±10） days, respectively. Survival in CTLA4Ig ＋ CD40LIg cotransfected group was significantly longer. Survivals of CTLA4Ig transfected group and CD40LIg transfected group were significantly longer than control group. In controJ animals, serum interleukin-2 and tumor necrosis factor a concentration significantly increased within seven days post transplantation. Haematoxylin eosin staining of grafts showed live islets in situ of transplant rats without inflammatory cell infiltration. Immunohistochemical staining confirmed the expression of insulin at islets in all experimental groups.Conclusions Transfer of CTLA4Ig and CD40Llg genes, especially the cotransfer of both, inhibits rejection of murine islet xenografts. Downregulated expressions of Th1 cells related cytokines mig
文摘At present, proven clinical treatments but no cures are available for diabetes, a global epidemic with a huge economic burden. Transplantation of islets ofLangerhans by their infusion into vascularized organs is an experimental clinical protocol, the first approach to attain cure. However, it is associated with lifelong use of immunosuppressants. To overcome the need for immunosuppression, islets are encapsulated and separated from the host immune system by a permselective membrane. The lead material for this application is alginate which was tested in many animal models and a few clinical trials. This review discusses all aspects related to the function of transplanted encapsulated islets such as the basic requirements from a permselective membrane(e.g., allowable hydrodynamic radii, implications of the thickness of the membrane and relative electrical charge). Another aspect involves adequate oxygen supply, which is essential for survival/performance of transplanted islets, especially when using large retrievable macrocapsules implanted in poorly oxygenated sites like the subcutis. Notably, islets can survive under low oxygen tension and are physiologically active at > 40 Torr. Surprisingly, when densely crowded, islets are fully functional under hyperoxic pressure of up to 500 Torr(> 300% of atmospheric oxygen tension). The review also addresses an additional category of requirements for optimal performance of transplanted islets, named auxiliary technologies. These include control of inflammation, apoptosis, angiogenesis, and the intra-capsular environment. The review highlights that curing diabetes with a functional bio-artificial pancreas requires optimizing all of these aspects, and that significant advances have already been made in many of them.
文摘Erdheim-Chester Disease (ECD) is a rare condition and has various differential diagnoses with other forms of histiocytosis, classified as one of non-Langerhans histiocytosis. The diagnosis of this condition remains challenging because its presentation includes non-specific systemic manifestations that can affect different organs caused by deposition of lipids and fibrosis. Most common include bone pain followed by progressive weakness and different lung manifestations. This case is about a rare presentation of ECD with Langerhans Histiocytosis as overlap syndrome, with findings of both diseases in a middle aged woman that presented dyspnea as the first symptom. The patient was treated initially as heart failure and remained without any improvement, being admitted to investigate. After a stricted follow-up, bone and lung involvement were noticed and a skin biopsy unveiled xanthomatized macrophages accompanied by Touton giant cells. This condition remains an important clinical entity and should provide new insights for clinicians dealing with respiratory diseases.
基金Supported by Hainan Academician Innovation Platform Fund,and the Hainan Province Clinical Medical Center.
文摘BACKGROUND Langerhans cell histiocytosis(LCH)is characterized by diabetes insipidus and is an uncommon occurrence.Pathological biopsies still have a certain degree of diagnostic probability.We present a case in which LCH initially affected the pituitary gland.This resulted in a misdiagnosis of chronic inflammation upon pathological examination.CASE SUMMARY A 25-year-old female exhibited symptoms of diabetes insipidus.Magnetic resonance imaging revealed an enhanced foci in the pituitary gland.After surgical resection of the pituitary lesion,the pathological diagnosis was chronic inflam-mation.However,the patient later experienced bone destruction in the skull and lower limb bones.After the lower limb bone lesion was compared with the initial pituitary lesion,the final diagnosis was modified to LCH.The patient was treated with multiple chemotherapy courses.However,the patient’s condition gradually worsened,and she eventually passed away at home.CONCLUSION LCH should be considered when patients exhibit diabetes insipidus and absence of high signal intensity in the pituitary gland on sagittal T1-weighted image and abnormal enhancement in the pituitary region.
基金the Chongqing Science and Health Committee Joint Medical Scientific Research General Project,No.2021MSXM236the Health Development Promotion Project,No.BJHA-CRP-086.
文摘BACKGROUND Langerhans cell histiocytosis(LCH)is a histiocytic proliferative disease caused by clonal proliferation of Langerhans cells,which is currently defined as an inflam-matory myeloid tumor.It is rare in adults,with an incidence of 1–2 per million,and is highly heterogeneous in clinical presentation,with unpredictable disease progression and outcome.CASE SUMMARY A 52-year-old postmenopausal female patient presented to the gynecology department in July 2023 with bilateral vulvar masses.She was diagnosed with recurrent multisystem LCH.The patient had previously been diagnosed with a single-system and single-focal LCH in October 2021 due to a right maxillofacial mass,which resolved after surgical treatment.A chemotherapy regimen was developed after multidisciplinary consultation.Six cycles of chemotherapy resulted in partial remission,and maintenance chemotherapy is currently being administered.CONCLUSION Recurrent LCH involving the bilateral vulva has been poorly reported.Compre-hensive imaging and pathological evaluation is important for diagnosis.The model of joint multidisciplinary specialist diagnosis and treatment is worthy of clinical application.
基金This work was supported by grants from the National Natural Science Foundation of China （No. 30972659）, Innovative Research Teams in Universities, Ministry of Education （No. IRT0760） and Liaoning Bureau （No. 2007T 186 and No. 2009T105）.
文摘Background The sensitization and elicitation phases are involved in the immunopathogenesis of contact hypersensitivity （CHS）. Langerhans cells （LCs） are believed to play pivotal roles in the sensitization stage of CHS. Local hyperthermia on skin induces the migration as well as maturation of epidermal LCs. Although fever-range whole body hyperthermia and local hyperthermia at 43℃ prior to sensitization were reported to suppress CHS, the effects of different temperatures and the timing sequence of local hyperthermia on CHS have not been tackled. Methods Local hyperthermia was applied to murine dorsal skin 3 days prior to, concurrent with, or 2 days post sensitization with fluorescein isothiocyanate （FITC） in BALB/c mice. Local hyperthermia temperatures at 37℃, 39℃, 41℃ and 43℃ were applied to mouse dorsal skin and the severity of CHS was calculated by measuring the swelling response of the challenged ears. Results Local hyperthermia at 39℃, 41℃ and 43℃ prior to sensitization reduced the severity of CHS, as compared with that at 37℃. The suppression of CHS was temperature dependant in that higher temperature had a stronger effect. On the contrary, the hyperthermia treatments, either concurrent with or post-sensitization, resulted in an enhanced temperature-dependant ear swelling response. Conclusions The severity of murine CHS could be influenced by local hyperthermia at the sensitization stage in a temperature dependant manner. The temporal effect of local hyperthermia suggested a novel factor in interpreting the severity of allergic contact dermatitis.
文摘Langerhans cell histiocytosis(LCH) is a rare syndrome characterized by unifocal,multifocal unisystem,or disseminated/multi-system disease that commonly involves the bone,skin,lymph nodes,pituitary,or sometimes lung(almost exclusively in smokers) causing a variety of symptoms from rashes and bone lesions to diabetes insipidus or pulmonary infiltrates.We present a previously unreported case of gastrointestinal LCH as well as a novel characteristic lesion affecting the colon of a young woman who presented with signs and symptoms mimicking acute on chronic appendicitis.Immunohistochemical analysis of appendectomy specimen and nodular specimens on colonoscopy demonstrated S-100,CD1a,and langerin reactivity.The patient underwent systemic chemotherapy with cytarabine and demonstrated excellent response to therapy.
基金This study was supported by grants from the National Science and Technology Key Projects （No. 2017ZX09304029004）, Beijing Municipal Science and Technology Commission （No. ZI71100001017050）, National Natural Science Foundation of China （No. 81700186）, Scientific Research Common Program of Beijing Municipal Commission of Education （No. KM201710025019）, Pediatric Project of Ai You Foundation （No. AYEK201802）, and Talent Training Project-Fostering Fund of National Natural Science Foundation of Beijing Children＇s Hospital, Capital Medical University （No. GPY201713）.
文摘Background： Pulmonary Langerhans cell histiocytosis （PLCH） is an interstitial primary pulmonary disease, characterized by Langerhans cell proliferation. It is easily misdiagnosed in children. This study aimed to characterize the clinical manifestations and features of PLCH by retrospective analysis. Meihods： A retrospective analysis was performed in 117 PLCH patients out of 338 LCH patients who were admitted in our center from November 2006 to October 2013. Variables between two groups were compared by Mann-Whitney U-test and Chi-square test. Kaplan-Meier curves were constructed to compare the survival rates and Cox regression to evaluate the effect of risk factors. Results： The median age of PLCH group was significantly lower than that ofnon-PkCH group （ 18.63 months vs. 43.4 months, P 〈 0.001 ）. All PLCH children had other organ involvement and only 11 cases （9.4%） had respiratory symptoms. The most common radiologic finding was cystic lesions （29 cases, 24.8%）. Pulmonary thnction abnormalities were dominated by obstructive ventilatory dysfunction （63 cases, 82.9%）. The 5-year overall survival （OS） of PECH children was 93.6% ± 2.3% and the event-free survival （EFS） was 55.7% ± 5.2%. Among the 38 cases with progressed or relapsed disease, five cases （13.2%） were due to progression or recurrence of lung damage. The 5-year OS of PECH children with ＂risk organ＂ involvement was significantly lower than those without ＂risk organ＂ involvement （86.0% ± 4.9% vs. 100%,X^2 = 8.793, P= 0.003）. The difference of EFS between two groups was also significant （43.7% ± 7.7% vs. 66.3% ± 6.5%,X^2 = 5.399, P = 0.020）. The ＂＇risk organ＂ involvement had a significant impact on survival （hazard ratio = 1.9, P = 0.039）. Conclusions： PLCH mainly occurs in young children, and only a small percentage of patients have respiratory symptoms. They generally have other organ involvement. Most of PLCH children have a good prognosis and most lung lesions could have improved
文摘Pulmonary Langerhans＇ cell histiocytosis （PLCH） is an isolated form of Langerhans＇ cell histiocytosis thatusually develops in cigarette smokers.1＇2^ PLCH usually has typical light microscopic morphology with a spectrum of progressive changes.2-4^ It is characterized by peribronchiolar Langerhans＇ cell infiltrates mixed with macrophages, lymphocytes and eosinophils, that form stellate nodules. The nodular lesions are frequently cavitated and form thick- and thin-walled cysts, which are thought to represent enlarged airway lumina. PLCH lesions display temporal microscopic heterogeneity, with a progression from dense cellular nodules to cavitary nodules with increasing degrees of fibrosis that may extend along alveolar walls. In advanced cases,
