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1.
Gastroenterology ; 158(1): 253-269.e14, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31593700

RESUMO

BACKGROUND & AIMS: Pancreatitis starts with primarily sterile local inflammation that induces systemic inflammatory response syndrome, followed by compensatory anti-inflammatory response syndrome (CARS). We investigated the mechanisms of these processes in mice and human serum. METHODS: We induced severe acute pancreatitis by partial duct ligation with caerulein stimulation or intraperitoneal injection of l-arginine in mice with deletion of interleukin (IL)12B, NLRP3, or IL18 and in mice given MCC950, a small molecule inhibitor of the NLRP3-inflammasome. Pancreata were collected from mice and analyzed by histology, and cytokine levels were measured in serum samples. We measured activation of adaptive immune responses in mice with pancreatitis by flow cytometry analysis of T cells (CD25 and CD69) isolated from the spleen. Differentiation of T-helper (Th1) cells, Th2 cells, and T-regulatory cells was determined by nuclear staining for TBET, GATA3, and FOXP3. We performed transcriptome analysis of mouse lymph nodes and bone marrow-derived macrophages after incubation with acini. We measured levels of cytokines in serum samples from patients with mild and severe acute pancreatitis. RESULTS: Activation of the adaptive immune response in mice was initiated by macrophage-derived, caspase 1-processed cytokines and required activation of NLRP3 (confirmed in serum samples from patients with pancreatitis). Spleen cells from mice with pancreatitis had increases in Th2 cells but not in Th1 cells. Bone marrow-derived macrophages secreted IL1B and IL18, but not IL12, after co-incubation with pancreatic acini. T-cell activation and severity of acute pancreatitis did not differ significantly between IL12B-deficient and control mice. In contrast, NLRP3- or IL18-deficient mice had reduced activation of T cells and no increase in Th2 cell-mediated responses compared with control mice. The systemic type 2 immune response was mediated by macrophage-derived cytokines of the IL1 family. Specifically, IL18 induced a Th2 cell-mediated response in the absence of IL12. MCC950 significantly reduced neutrophil infiltration, T-cell activation, and disease severity in mice. CONCLUSIONS: In mice with severe pancreatitis, we found systemic inflammatory response syndrome and compensatory anti-inflammatory response syndrome developed in parallel. Infiltrating macrophages promote inflammation and simultaneously induce a Th2 cell-mediated response via IL18. Inhibition of NLRP3 reduces systemic inflammatory response syndrome and compensatory anti-inflammatory response syndrome and might be used to treat patients with severe pancreatitis.


Assuntos
Furanos/administração & dosagem , Inflamassomos/imunologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Pancreatite/imunologia , Sulfonamidas/administração & dosagem , Síndrome de Resposta Inflamatória Sistêmica/imunologia , Células Acinares , Imunidade Adaptativa , Animais , Arginina/toxicidade , Células Cultivadas , Ceruletídeo/toxicidade , Citocinas/sangue , Citocinas/imunologia , Modelos Animais de Doenças , Humanos , Injeções Intraperitoneais , Interleucina-18/imunologia , Interleucina-18/metabolismo , Macrófagos/imunologia , Camundongos , Camundongos Knockout , Proteína 3 que Contém Domínio de Pirina da Família NLR/antagonistas & inibidores , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Pâncreas/citologia , Pâncreas/imunologia , Pâncreas/patologia , Pancreatite/induzido quimicamente , Pancreatite/tratamento farmacológico , Cultura Primária de Células , Síndrome de Resposta Inflamatória Sistêmica/sangue , Síndrome de Resposta Inflamatória Sistêmica/tratamento farmacológico , Células Th2/imunologia , Células Th2/metabolismo
2.
Genes Dev ; 33(21-22): 1475-1490, 2019 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-31676735

RESUMO

A comprehensive understanding of mechanisms that underlie the development and function of human cells requires human cell models. For the pancreatic lineage, protocols have been developed to differentiate human pluripotent stem cells (hPSCs) into pancreatic endocrine and exocrine cells through intermediates resembling in vivo development. In recent years, this differentiation system has been employed to decipher mechanisms of pancreatic development, congenital defects of the pancreas, as well as genetic forms of diabetes and exocrine diseases. In this review, we summarize recent insights gained from studies of pancreatic hPSC models. We discuss how genome-scale analyses of the differentiation system have helped elucidate roles of chromatin state, transcription factors, and noncoding RNAs in pancreatic development and how the analysis of cells with disease-relevant mutations has provided insight into the molecular underpinnings of genetically determined diseases of the pancreas.


Assuntos
Modelos Biológicos , Pâncreas/citologia , Pâncreas/crescimento & desenvolvimento , Células-Tronco Pluripotentes/citologia , Diferenciação Celular , Estudo de Associação Genômica Ampla , Humanos , Pâncreas/patologia , Pancreatopatias/genética , Pancreatopatias/fisiopatologia , Células-Tronco Pluripotentes/fisiologia
3.
Pathologe ; 40(Suppl 3): 311-315, 2019 Dec.
Artigo em Alemão | MEDLINE | ID: mdl-31705231

RESUMO

BACKGROUND: Cytology has a key role in the step-wise diagnostic approach to pancreatic mass lesions. Brush cytology and ultrasound-guided endoscopic fine-needle aspiration provide specimens for diagnosis prior to surgical or conservative therapy. The diagnostic system of the Papanicolaou Society of Cytopathology provides a conceptual framework for reporting these specimens. Cystic lesions represent a particular challenge in pancreatic cytology, as in many instances a purely morphological approach will not result in an adequate diagnostic interpretation. Noteworthy from a conceptual point of view is how the Papanicolaou Society System incorporates non-morphological methods: laboratory chemical (CEA >192 ng/ml) and molecular (KRAS and/or GNAS mutations) findings are part of the formal diagnostic criteria for neoplastic cysts. RESULTS: The Bern experience shows that such an integrated approach results in a significantly increased diagnostic yield. Among 83 samples analyzed, adequate DNA could be extracted in 79 samples (95%). Next generation sequencing identified pathogenic mutations in 46 cases (58%). Of these, in 35 (76%) a neoplastic cyst could not have been diagnosed by morphology alone. CONCLUSION: These findings illustrate a new perspective for diagnostic situations, where morphology alone does allow for a sufficient diagnostic work-up. Along this line of thinking, liquid biopsy should not be regarded as a replacement, but rather an extension of the cytology's diagnostic armamentarium, according to the principle of "doing more with less."


Assuntos
Pâncreas/citologia , Cisto Pancreático , Neoplasias Pancreáticas , Análise Mutacional de DNA , DNA de Neoplasias/genética , Aspiração por Agulha Fina Guiada por Ultrassom Endoscópico , Humanos , Mutação , Cisto Pancreático/diagnóstico , Cisto Pancreático/genética , Cisto Pancreático/patologia , Neoplasias Pancreáticas/diagnóstico , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia
4.
Nat Commun ; 10(1): 4517, 2019 10 04.
Artigo em Inglês | MEDLINE | ID: mdl-31586069

RESUMO

Neonatal inflammatory diseases are associated with severe morbidity, but the inflammatory factors underlying them and their potential effector mechanisms are poorly defined. Here we show that necrotizing enterocolitis in neonate mice is accompanied by elevation of IL-23 and IL-22 and decreased production of pancreatic enzymes. These phenotypes are mirrored in neonate mice overexpressing IL-23 in CX3CR1+ myeloid cells or in keratinocytes. The mice fail to grow and die prematurely, displaying systemic inflammation, nutrient malabsorption and decreased expression of intestinal and pancreatic genes mediating digestion and absorption of carbohydrates, proteins, and lipids. Germ-free environment improves, and genetic ablation of IL-22 restores normal growth in mice overexpressing IL-23. Mechanistically, IL-22 acts directly at the level of pancreatic acinar cells to decrease expression of the pancreas associated transcription factor 1a (PTF1a). These results show that augmented production of IL-23 and IL-22 in early life has a negative impact on pancreatic enzyme secretion and food absorption.


Assuntos
Enterocolite Necrosante/imunologia , Interleucina-23/metabolismo , Interleucinas/metabolismo , Pâncreas/enzimologia , Fatores de Transcrição/metabolismo , Células Acinares/enzimologia , Animais , Animais Recém-Nascidos , Células Cultivadas , Modelos Animais de Doenças , Enterocolite Necrosante/patologia , Humanos , Interleucina-23/genética , Interleucina-23/imunologia , Interleucinas/genética , Interleucinas/imunologia , Absorção Intestinal/imunologia , Mucosa Intestinal/imunologia , Mucosa Intestinal/patologia , Queratinócitos , Camundongos , Camundongos Knockout , Células Mieloides , Pâncreas/citologia , Cultura Primária de Células
5.
Nature ; 574(7776): 112-116, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31554966

RESUMO

Organogenesis is a complex and interconnected process that is orchestrated by multiple boundary tissue interactions1-7. However, it remains unclear how individual, neighbouring components coordinate to establish an integral multi-organ structure. Here we report the continuous patterning and dynamic morphogenesis of hepatic, biliary and pancreatic structures, invaginating from a three-dimensional culture of human pluripotent stem cells. The boundary interactions between anterior and posterior gut spheroids differentiated from human pluripotent stem cells enables retinoic acid-dependent emergence of hepato-biliary-pancreatic organ domains specified at the foregut-midgut boundary organoids in the absence of extrinsic factors. Whereas transplant-derived tissues are dominated by midgut derivatives, long-term-cultured microdissected hepato-biliary-pancreatic organoids develop into segregated multi-organ anlages, which then recapitulate early morphogenetic events including the invagination and branching of three different and interconnected organ structures, reminiscent of tissues derived from mouse explanted foregut-midgut culture. Mis-segregation of multi-organ domains caused by a genetic mutation in HES1 abolishes the biliary specification potential in culture, as seen in vivo8,9. In sum, we demonstrate that the experimental multi-organ integrated model can be established by the juxtapositioning of foregut and midgut tissues, and potentially serves as a tractable, manipulatable and easily accessible model for the study of complex human endoderm organogenesis.


Assuntos
Sistema Biliar/embriologia , Intestinos/embriologia , Fígado/embriologia , Modelos Biológicos , Morfogênese , Pâncreas/embriologia , Animais , Sistema Biliar/citologia , Biomarcadores/análise , Biomarcadores/metabolismo , Padronização Corporal , Endoderma/citologia , Endoderma/embriologia , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Intestinos/citologia , Fígado/citologia , Masculino , Camundongos , Organoides/citologia , Organoides/embriologia , Pâncreas/citologia , Esferoides Celulares/citologia , Esferoides Celulares/metabolismo , Esferoides Celulares/transplante , Fatores de Transcrição HES-1/análise , Fatores de Transcrição HES-1/metabolismo
6.
World J Gastroenterol ; 25(32): 4673-4681, 2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31528093

RESUMO

Zollinger-Ellison syndrome (ZES) is characterized by gastric acid hypersecretion causing severe recurrent acid-related peptic disease. Excessive secretion of gastrin can now be effectively controlled with powerful proton pump inhibitors, but surgical management to control gastrinoma itself remains controversial. Based on a thorough literature review, we design a surgical algorithm for ZES and list some significant consensus findings and recommendations: (1) For sporadic ZES, surgery should be routinely undertaken as early as possible not only for patients with a precisely localized diagnosis but also for those with negative imaging findings. The surgical approach for sporadic ZES depends on the lesion location (including the duodenum, pancreas, lymph nodes, hepatobiliary tract, stomach, and some extremely rare sites such as the ovaries, heart, omentum, and jejunum). Intraoperative liver exploration and lymphadenectomy should be routinely performed; (2) For multiple endocrine neoplasia type 1-related ZES (MEN1/ZES), surgery should not be performed routinely except for lesions > 2 cm. An attempt to perform radical resection (pancreaticoduodenectomy followed by lymphadenectomy) can be made. The ameliorating effect of parathyroid surgery should be considered, and parathyroidectomy should be performed first before any abdominal surgery for ZES; and (3) For hepatic metastatic disease, hepatic resection should be routinely performed. Currently, liver transplantation is still considered an investigational therapeutic approach for ZES. Well-designed prospective studies are desperately needed to further verify and modify the current considerations.


Assuntos
Gastroenterologia/normas , Oncologia/normas , Guias de Prática Clínica como Assunto , Síndrome de Zollinger-Ellison/cirurgia , Duodeno/citologia , Duodeno/patologia , Duodeno/cirurgia , Células Secretoras de Gastrina/patologia , Gastrinas/metabolismo , Gastroenterologia/métodos , Hepatectomia , Humanos , Fígado/citologia , Fígado/patologia , Fígado/cirurgia , Excisão de Linfonodo , Oncologia/métodos , Pâncreas/citologia , Pâncreas/patologia , Pâncreas/cirurgia , Pancreaticoduodenectomia , Paratireoidectomia , Estômago/citologia , Estômago/patologia , Estômago/cirurgia , Fatores de Tempo , Síndrome de Zollinger-Ellison/patologia
7.
Gastroenterology ; 157(6): 1660-1672.e2, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31493399

RESUMO

BACKGROUND & AIMS: Pancreatitis is characterized by increased influx of Ca2+ into acinar cells, by unknown mechanisms. Inhibitors of Ca2+ influx channels could be effective in treating acute pancreatitis, but these have deleterious side effects that can result in death. We investigated the expression patterns and functions of acinar cell Ca2+ channels and factors that regulate them during development of acute pancreatitis, along with changes in the channel inactivator store-operated calcium entry-associated regulatory factor (SARAF). We investigated whether SARAF is a target for treatment of acute pancreatitis and its status in human with pancreatitis. METHODS: We generated mice that expressed SARAF tagged with hemagglutinin, using CRISPR/Cas9 gene editing, and isolated acinar cells. We also performed studies with Saraf-/- mice, Sarafzf/zf mice, mice without disruption of Saraf (control mice), and mice that overexpress fluorescently labeled SARAF in acinar cells. We analyzed interactions between stromal interaction molecule 1 (STIM1) and SARAF in HEK cells stimulated with carbachol using fluorescence resonance energy transfer microscopy and immunoprecipitation. Mice were given injections of caerulein or L-arginine to induce pancreatitis. Pancreatic tissues and blood samples were collected and levels of serum amylase, trypsin, tissue damage, inflammatory mediators, and inflammatory cells were measured. We performed quantitative polymerase chain reaction analyses of pancreatic tissues from 6 organ donors without pancreatic disease (controls) and 8 patients with alcohol-associated pancreatitis. RESULTS: Pancreatic levels of Ca2+ influx channels or STIM1 did not differ significantly between acinar cells from mice with vs. without pancreatitis. By contrast, pancreatic levels of Saraf messenger RNA and SARAF protein initially markedly increased but then decreased during cell stimulation or injection of mice with caerulein, resulting in excessive Ca2+ influx. STIM1 interacted stably with SARAF following stimulation of HEK or mouse acinar cells with physiologic levels of carbachol, but only transiently following stimulation with pathologic levels of carbachol, leading to excessive Ca2+ influx. We observed reduced levels of SARAF messenger RNA in pancreatic tissues from patients with pancreatitis, compared with controls. SARAF knockout mice developed more severe pancreatitis than control mice after administration of caerulein or L-arginine, and pancreatic acinar cells from these mice had significant increases in Ca2+ influx. Conversely, overexpression of SARAF in acini reduced Ca2+ influx, eliminated inflammation, and reduced severity of acute pancreatitis. CONCLUSIONS: In mice with pancreatitis, SARAF initially increases but is then degraded, resulting in excessive, pathological Ca2+ influx by acinar cells. SARAF knockout mice develop more severe pancreatitis than control mice, whereas mice that express SARAF from a transgene in acinar cells develop less-severe pancreatitis. SARAF therefore appears to prevent pancreatic damage during development of acute pancreatitis. Strategies to stabilize or restore SARAF to acinar cells might be developed for treatment of pancreatitis.


Assuntos
Cálcio/metabolismo , Proteínas Sensoras de Cálcio Intracelular/metabolismo , Proteínas de Membrana/metabolismo , Proteínas de Neoplasias/metabolismo , Pâncreas/patologia , Pancreatite/patologia , Molécula 1 de Interação Estromal/metabolismo , Células Acinares/patologia , Animais , Ceruletídeo/toxicidade , Modelos Animais de Doenças , Células HEK293 , Humanos , Proteínas Sensoras de Cálcio Intracelular/genética , Proteínas de Membrana/genética , Camundongos , Camundongos Knockout , Pâncreas/citologia , Pancreatite/sangue , Pancreatite/induzido quimicamente , Índice de Gravidade de Doença
8.
Int J Mol Sci ; 20(17)2019 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-31438545

RESUMO

Pancreatic progenitor cells (PPCs) are the primary source for all pancreatic cells, including beta-cells, and thus the proliferation and differentiation of PPCs into islet-like cell clusters (ICCs) opens an avenue to providing transplantable islets for diabetic patients. Meanwhile, mesenchymal stem cells (MSCs) can enhance the development and function of different cell types of interest, but their role on PPCs remains unknown. We aimed to explore the mechanism-of-action whereby MSCs induce the in vitro and in vivo PPC/ICC development by means of our established co-culture system of human PPCs with human fetal bone marrow-derived MSCs. We examined the effect of MSC-conditioned medium on PPC proliferation and survival. Meanwhile, we studied the effect of MSC co-culture enhanced PPC/ICC function in vitro and in vivo co-/transplantation. Furthermore, we identified IGF1 as a critical factor responsible for the MSC effects on PPC differentiation and proliferation via IGF1-PI3K/Akt and IGF1-MEK/ERK1/2, respectively. In conclusion, our data indicate that MSCs stimulated the differentiation and proliferation of human PPCs via IGF1 signaling, and more importantly, promoted the in vivo engraftment function of ICCs. Taken together, our protocol may provide a mechanism-driven basis for the proliferation and differentiation of PPCs into clinically transplantable islets.


Assuntos
Diferenciação Celular/fisiologia , Proliferação de Células/fisiologia , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/fisiologia , Pâncreas/citologia , Células-Tronco/citologia , Células-Tronco/fisiologia , Apoptose/fisiologia , Células Cultivadas , Técnicas de Cocultura , Humanos , Células-Tronco Mesenquimais/citologia , Fosfatidilinositol 3-Quinases/metabolismo , Células-Tronco/metabolismo
9.
Genome Biol ; 20(1): 165, 2019 08 12.
Artigo em Inglês | MEDLINE | ID: mdl-31405383

RESUMO

To fully utilize the power of single-cell RNA sequencing (scRNA-seq) technologies for identifying cell lineages and bona fide transcriptional signals, it is necessary to combine data from multiple experiments. We present BERMUDA (Batch Effect ReMoval Using Deep Autoencoders), a novel transfer-learning-based method for batch effect correction in scRNA-seq data. BERMUDA effectively combines different batches of scRNA-seq data with vastly different cell population compositions and amplifies biological signals by transferring information among batches. We demonstrate that BERMUDA outperforms existing methods for removing batch effects and distinguishing cell types in multiple simulated and real scRNA-seq datasets.


Assuntos
Aprendizado Profundo , Perfilação da Expressão Gênica/métodos , Análise de Sequência de RNA/métodos , Algoritmos , Humanos , Leucócitos Mononucleares/metabolismo , Pâncreas/citologia , Pâncreas/metabolismo , Análise de Célula Única/métodos , Linfócitos T/metabolismo
11.
PLoS Biol ; 17(7): e3000382, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31323030

RESUMO

The Hippo pathway directs cell differentiation during organogenesis, in part by restricting proliferation. How Hippo signaling maintains a proliferation-differentiation balance in developing tissues via distinct molecular targets is only beginning to be understood. Our study makes the unexpected finding that Hippo suppresses nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB) signaling in pancreatic progenitors to permit cell differentiation and epithelial morphogenesis. We find that pancreas-specific deletion of the large tumor suppressor kinases 1 and 2 (Lats1/2PanKO) from mouse progenitor epithelia results in failure to differentiate key pancreatic lineages: acinar, ductal, and endocrine. We carried out an unbiased transcriptome analysis to query differentiation defects in Lats1/2PanKO. This analysis revealed increased expression of NFκB activators, including the pantetheinase vanin1 (Vnn1). Using in vivo and ex vivo studies, we show that VNN1 activates a detrimental cascade of processes in Lats1/2PanKO epithelium, including (1) NFκB activation and (2) aberrant initiation of epithelial-mesenchymal transition (EMT), which together disrupt normal differentiation. We show that exogenous stimulation of VNN1 or NFκB can trigger this cascade in wild-type (WT) pancreatic progenitors. These findings reveal an unexpected requirement for active suppression of NFκB by LATS1/2 during pancreas development, which restrains a cell-autonomous deleterious transcriptional program and thereby allows epithelial differentiation.


Assuntos
Diferenciação Celular/genética , Transição Epitelial-Mesenquimal/genética , NF-kappa B/genética , Pâncreas/metabolismo , Proteínas Serina-Treonina Quinases/genética , Células-Tronco/metabolismo , Proteínas Supressoras de Tumor/genética , Animais , Proliferação de Células/genética , Perfilação da Expressão Gênica/métodos , Camundongos Knockout , Camundongos Transgênicos , Microscopia Confocal , NF-kappa B/metabolismo , Pâncreas/citologia , Pâncreas/embriologia , Proteínas Serina-Treonina Quinases/metabolismo , Técnicas de Cultura de Tecidos , Proteínas Supressoras de Tumor/metabolismo
12.
Indian J Pathol Microbiol ; 62(3): 477-480, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31361246

RESUMO

We present an autopsy case of a 19 year old male admitted for breathlessness and oliguria. He was diabetic since 7 years of age and was on insulin. Patient was on testosterone and anti hypertensives. He was diagnosed of hypocontractile bladder and congenital bilateral megaureter with vesico-ureteric reflux 2 years back. History of hemiparesis 2 years back. CT scan of the brain showed a right fronto- parietal healed infarct. At autopsy, bilateral kidneys showed coarse granularity and scarring. Pelvicalyceal system and both ureters were dilated. A right sided intrabdominal testes was identified. On histology, kidney showed features of diabetic nephropathy and pancreas showed decreased number of islet cells. Correlating the clinical, laboratory and autopsy parameters, our case satisfies the EURO-WABB criteria (1major+2minor) for diagnosis of Wolfram Syndrome, even though genetic confirmation could not be done.[1],[2].


Assuntos
Encéfalo/patologia , Síndrome de Wolfram/diagnóstico por imagem , Autopsia , Encéfalo/diagnóstico por imagem , Complicações do Diabetes , Nefropatias Diabéticas , Evolução Fatal , Humanos , Rim/patologia , Masculino , Pâncreas/citologia , Pâncreas/patologia , Paresia , Bexiga Urinária/patologia , Adulto Jovem
13.
Soft Matter ; 15(24): 4827-4835, 2019 Jun 19.
Artigo em Inglês | MEDLINE | ID: mdl-31180412

RESUMO

Self-assembling peptides constitute an important class of functional biomaterials. A number of short amyloidogenic stretches have been identified from amyloid proteins. Such peptides, as such or through subtle modifications, can turn out to be promising candidates for functional biomaterials. End-capped Aß16-22, the well-studied amyloidogenic stretch from ß-amyloid, is reported to be non-hydrogelating up to 20 mM concentration. Here we investigated the hydrogelation propensity of Aß16-22 repeats connected through ß-turn-supporting motifs. The peptide repeats connected through Asn-Gly, Aib-DPro, and DPro-Gly formed transparent hydrogels at concentrations ≥2 mM. The repeats of the aromatic analog Aß16-22(F20Y) also resulted in similar hydrogels. Like other peptide-based gels reported earlier, these gels could trap the anticancer drug doxorubicin and displayed steady release in water. In addition, the gels supported the growth of mammalian cell lines, HEK-293 and RIN-5F. These data show that turn-inducing motifs can have marked effects on the hydrogelating propensity of self-assembling peptides.


Assuntos
Peptídeos beta-Amiloides/química , Doxorrubicina/farmacocinética , Portadores de Fármacos/farmacocinética , Hidrogéis/química , Fragmentos de Peptídeos/química , Motivos de Aminoácidos , Animais , Técnicas de Cultura de Células/instrumentação , Técnicas de Cultura de Células/métodos , Dicroísmo Circular , Portadores de Fármacos/química , Células HEK293 , Humanos , Insulina/genética , Microscopia de Força Atômica , Pâncreas/citologia , Ratos , Sequências Repetitivas de Aminoácidos , Espectroscopia de Infravermelho com Transformada de Fourier
14.
Acta Histochem ; 121(5): 638-645, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31146895

RESUMO

In the human pancreas, various forms of endocrine cell arrangement are found: single endocrine cells, endocrine cell clusters, and mantel, bipolar and mosaic cell (mixed) islets. Our aim was to analyse the distribution and dynamics of insulin-, glucagon- and somatostatin-containing cells within the various forms of endocrine pancreas arrangement during human prenatal development and in adults and to suggest a mechanism of change in the endocrine cell ratio in adult islets. Pancreatic autopsies derived from human foetuses from the 10th to the 40th weeks of development and from adults were examined using histological, immunohistochemical and morphometric methods. During development, the human endocrine pancreas undergoes not only de novo differentiation of endocrine cells and islet formation, but morphogenetic restructuring, which is revealed as a change of the α-, ß- and δ-cell ratio in the islets. In particular, increased proportion of glucagon- and somatostatin-containing cells and decreased proportion of ß-cells were shown in the largest mosaic islets in adults. Our results indicate that the distribution and proportion of α-, ß- and δ-cells depend on the islets size and vascularisation. Studying of the mechanism of such restructuring may contribute to the development of new approaches in the treatment of diabetes mellitus.


Assuntos
Células Secretoras de Glucagon/citologia , Células Secretoras de Insulina/citologia , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/embriologia , Pâncreas/citologia , Células Secretoras de Somatostatina/citologia , Desenvolvimento Embrionário , Humanos
15.
In Vitro Cell Dev Biol Anim ; 55(6): 453-461, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31140102

RESUMO

Although bone marrow-derived mesenchymal stem cells (BMSCs) have been reported to be effective for the attenuation of diabetes, they have limitations. Whether BMSCs can be target-induced by pancreatic stem cells (PSCs) to have effectiveness for the restoration of diabetic islet injury was unknown. In this study, based on their successful isolation and cultivation, BMSCs were co-cultured with PSCs. The pancreatic stem cells markers, Nestin and Neurogenin3 in co-cultured BMSCs were detected to evaluate the target-induction effects. After the diabetic rats were intravenously injected with the target-induced BMSCs, general indicators and islet morphology were detected. The islet insulin generation, and serum insulin and C-peptide contents were measured. It was found that after co-culture, the mRNA expressions, protein contents and distributions of Nestin and Neurogenin3, were dramatically high in BMSCs, indicating that they were successfully target-induced to pancreatic stem-like cells. Furthermore, the target-induced BMSCs had beneficial effects on serum glycated albumin levels and glycogen contents as well as islet morphology of the diabetic rats. Besides elevation of islet insulin generation, the target-induced BMSCs had significant effect on serum insulin and C-peptide contents. In conclusion, BMSCs could be target-induced by PSCs to have effectiveness on the pancreatic restoration of diabetic rats.


Assuntos
Diabetes Mellitus Experimental/terapia , Ilhotas Pancreáticas/patologia , Transplante de Células-Tronco Mesenquimais/métodos , Células-Tronco Mesenquimais/fisiologia , Pâncreas/citologia , Animais , Animais Recém-Nascidos , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Células da Medula Óssea/citologia , Peptídeo C/metabolismo , Técnicas de Cocultura , Diabetes Mellitus Experimental/patologia , Glicogênio/metabolismo , Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Masculino , Células-Tronco Mesenquimais/citologia , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Nestina/genética , Nestina/metabolismo , Ratos Sprague-Dawley , Albumina Sérica/análise , Albumina Sérica/metabolismo , Células-Tronco/citologia
16.
Life Sci ; 226: 57-67, 2019 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-30930115

RESUMO

AIM: At performing a temporal analysis of the distribution pattern of islet endocrine cells and antioxidant enzymes in diabetic rats during the post-natal critical development window. MAIN METHODS: The newborns received streptozotocin (STZ) at birth for diabetes induction, and control females received the vehicle. The animals were euthanized at different lifetimes: D5, D10, D15, and D30. Morphological analysis of pancreas and biochemical assays was performed. KEY FINDINGS: The STZ-induced rats presented irregular shape of islet on D5 and there was an attempt to restore of this shape in other life moment studied. There was an increase progressive in islet area, however they maintained smaller than those of control rats, with lower labeling intensity for insulin, higher for glucagon and somatostatin, lower for SOD-1 was lower in the islets of the STZ-induced animals at all times studied and for GSH-Px in D10 and D30. SIGNIFICANCE: Although STZ-induced diabetic rats presented compensatory mechanisms to restore the mass of endocrine cells, this was not sufficient since these rats developed the diabetic state. This was confirmed by the oral glucose tolerance test from D30. In addition, the delta (δ)-cells presented ectopic location in islets, indicating a possible relationship for beta (ß)-cell mass restoration. There was a response of the pancreas to reduce the hyperglycemia in the first month of life. Furthermore, the cells from the endocrine pancreas of diabetic animals show a decline of antioxidant enzymatic, contributing to the increased susceptibility of cells to hyperglycemia-induced ROS in this postnatal critical development window.


Assuntos
Diabetes Mellitus Experimental/metabolismo , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/metabolismo , Animais , Antioxidantes/análise , Antioxidantes/metabolismo , Glicemia/metabolismo , Diabetes Mellitus Experimental/induzido quimicamente , Feminino , Glucagon , Glucose/metabolismo , Hiperglicemia , Insulina , Células Secretoras de Insulina , Masculino , Pâncreas/citologia , Gravidez , Ratos , Ratos Wistar , Análise Espaço-Temporal , Estreptozocina/farmacologia
17.
Int J Mol Sci ; 20(7)2019 Apr 05.
Artigo em Inglês | MEDLINE | ID: mdl-30959771

RESUMO

Mitochondrial dysfunction is a core feature of acute pancreatitis, a severe disease in which oxidative stress is elevated. Mitochondrial targeting of antioxidants is a potential therapeutic strategy for this and other diseases, although thus far mixed results have been reported. We investigated the effects of mitochondrial targeting with the antioxidant MitoQ on pancreatic acinar cell bioenergetics, adenosine triphosphate (ATP) production and cell fate, in comparison with the non-antioxidant control decyltriphenylphosphonium bromide (DecylTPP) and general antioxidant N-acetylcysteine (NAC). MitoQ (µM range) and NAC (mM range) caused sustained elevations of basal respiration and the inhibition of spare respiratory capacity, which was attributable to an antioxidant action since these effects were minimal with DecylTPP. Although MitoQ but not DecylTPP decreased cellular NADH levels, mitochondrial ATP turnover capacity and cellular ATP concentrations were markedly reduced by both MitoQ and DecylTPP, indicating a non-specific effect of mitochondrial targeting. All three compounds were associated with a compensatory elevation of glycolysis and concentration-dependent increases in acinar cell apoptosis and necrosis. These data suggest that reactive oxygen species (ROS) contribute a significant negative feedback control of basal cellular metabolism. Mitochondrial targeting using positively charged molecules that insert into the inner mitochondrial member appears to be deleterious in pancreatic acinar cells, as does an antioxidant strategy for the treatment of acute pancreatitis.


Assuntos
Células Acinares/metabolismo , Antioxidantes/metabolismo , Linhagem da Célula , Metabolismo Energético , Mitocôndrias/metabolismo , Pâncreas/citologia , Acetilcisteína/farmacologia , Células Acinares/efeitos dos fármacos , Trifosfato de Adenosina/biossíntese , Animais , Morte Celular/efeitos dos fármacos , Linhagem da Célula/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Flavina-Adenina Dinucleotídeo/metabolismo , Camundongos Endogâmicos C57BL , Mitocôndrias/efeitos dos fármacos , NAD/metabolismo , Oniocompostos/farmacologia , Compostos Organofosforados/farmacologia , Oxirredução , Ubiquinona/análogos & derivados , Ubiquinona/farmacologia
18.
Pac Symp Biocomput ; 24: 350-361, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30963074

RESUMO

Single-cell RNA sequencing (scRNA-seq) techniques have been very powerful in analyzing heterogeneous cell population and identifying cell types. Visualizing scRNA-seq data can help researchers effectively extract meaningful biological information and make new discoveries. While commonly used scRNA-seq visualization methods, such as t-SNE, are useful in detecting cell clusters, they often tear apart the intrinsic continuous structure in gene expression profiles. Topological Data Analysis (TDA) approaches like Mapper capture the shape of data by representing data as topological networks. TDA approaches are robust to noise and different platforms, while preserving the locality and data continuity. Moreover, instead of analyzing the whole dataset, Mapper allows researchers to explore biological meanings of specific pathways and genes by using different filter functions. In this paper, we applied Mapper to visualize scRNA-seq data. Our method can not only capture the clustering structure of cells, but also preserve the continuous gene expression topologies of cells. We demonstrated that by combining with gene co-expression network analysis, our method can reveal differential expression patterns of gene co-expression modules along the Mapper visualization.


Assuntos
RNA/genética , Análise de Sequência de RNA/estatística & dados numéricos , Análise de Célula Única/estatística & dados numéricos , Algoritmos , Biologia Computacional , Interpretação Estatística de Dados , Bases de Dados Genéticas/estatística & dados numéricos , Perfilação da Expressão Gênica/estatística & dados numéricos , Redes Reguladoras de Genes , Humanos , Melanoma/genética , Pâncreas/citologia , Pâncreas/metabolismo
19.
Int J Mol Sci ; 20(8)2019 Apr 22.
Artigo em Inglês | MEDLINE | ID: mdl-31013667

RESUMO

UbiA prenyltransferase domain-containing protein 1 (UBIAD1) is a vitamin K2 biosynthetic enzyme. We previously showed the lethality of this enzyme in UBIAD1 knockout mice during the embryonic stage. However, the biological effects of UBIAD1 deficiency after birth remain unclear. In the present study, we used a tamoxifen-inducible systemic UBIAD1 knockout mouse model to determine the role of UBIAD1 in adult mice. UBIAD1 knockout resulted in the death of the mice within about 60 days of administration of tamoxifen. The pancreas presented with the most prominent abnormality in the tamoxifen-induced UBIAD1 knockout mice. The pancreas was reduced remarkably in size; furthermore, the pancreatic acinar cells disappeared and were replaced by vacuoles. Further analysis revealed that the vacuoles were adipocytes. UBIAD1 deficiency in the pancreatic acinar cells caused an increase in oxidative stress and autophagy, leading to apoptotic cell death in the tamoxifen-induced UBIAD 1 knockout mice. These results indicate that UBIAD1 is essential for maintaining the survival of pancreatic acinar cells in the pancreas.


Assuntos
Células Acinares/metabolismo , Dimetilaliltranstransferase/genética , Pâncreas/citologia , Pâncreas/metabolismo , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Animais , Apoptose/efeitos dos fármacos , Atrofia , Autofagia/efeitos dos fármacos , Linhagem Celular , Sobrevivência Celular/genética , Dimetilaliltranstransferase/metabolismo , Feminino , Genes Letais , Genótipo , Imuno-Histoquímica , Células-Tronco Mesenquimais/metabolismo , Camundongos , Camundongos Knockout , Infiltração de Neutrófilos , Neutrófilos/metabolismo , Neutrófilos/patologia , Estresse Oxidativo/efeitos dos fármacos , Pâncreas/patologia , Fenótipo , Tamoxifeno/farmacologia
20.
PLoS One ; 14(4): e0215137, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30973910

RESUMO

Hybrid 3D scaffolds composed of different biomaterials with fibrous structure or enriched with different inclusions (i.e., nano- and microparticles) have already demonstrated their positive effect on cell integration and regeneration. The analysis of fibers in hybrid biomaterials, especially in a 3D space is often difficult due to their various diameters (from micro to nanoscale) and compositions. Though biomaterials processing workflows are implemented, there are no software tools for fiber analysis that can be easily integrated into such workflows. Due to the demand for reproducible science with Jupyter notebooks and the broad use of the Python programming language, we have developed the new Python package quanfima offering a complete analysis of hybrid biomaterials, that include the determination of fiber orientation, fiber and/or particle diameter and porosity. Here, we evaluate the provided tensor-based approach on a range of generated datasets under various noise conditions. Also, we show its application to the X-ray tomography datasets of polycaprolactone fibrous scaffolds pure and containing silicate-substituted hydroxyapatite microparticles, hydrogels enriched with bioglass contained strontium and alpha-tricalcium phosphate microparticles for bone tissue engineering and porous cryogel 3D scaffold for pancreatic cell culturing. The results obtained with the help of the developed package demonstrated high accuracy and performance of orientation, fibers and microparticles diameter and porosity analysis.


Assuntos
Materiais Biocompatíveis/química , Regeneração Óssea , Pâncreas/citologia , Software , Engenharia Tecidual/métodos , Tecidos Suporte , Automação , Células Cultivadas , Humanos , Modelos Biológicos , Poliésteres/química
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