Heterogeneity of Diabetes: ß-Cells, Phenotypes, and Precision Medicine: Proceedings of an International Symposium of the Canadian Institutes of Health Research's Institute of Nutrition, Metabolism and Diabetes and the U.S. National Institutes of Health's National Institute of Diabetes and Digestive and Kidney Diseases.

One hundred years have passed since the discovery of insulin-an achievement that transformed diabetes from a fatal illness into a manageable chronic condition. The decades since that momentous achievement have brought ever more rapid innovation and advancement in diabetes research and clinical care. To celebrate the important work of the past century and help to chart a course for its continuation into the next, the Canadian Institutes of Health Research's Institute of Nutrition, Metabolism and Diabetes and the U.S. National Institutes of Health's National Institute of Diabetes and Digestive and Kidney Diseases recently held a joint international symposium, bringing together a cohort of researchers with diverse interests and backgrounds from both countries and beyond to discuss their collective quest to better understand the heterogeneity of diabetes and thus gain insights to inform new directions in diabetes treatment and prevention. This article summarizes the proceedings of that symposium, which spanned cutting-edge research into various aspects of islet biology, the heterogeneity of diabetic phenotypes, and the current state of and future prospects for precision medicine in diabetes.

Measurement of proliferative responses of cultured lymphocytes.

Measurement of proliferative responses of human lymphocytes is a fundamental technique for the assessment of their biological responses to various stimuli. Most simply, this involves measurement of the number of cells present in a culture before and after the addition of a stimulating agent. This unit contains several different prototype protocols to induce proliferation in lymphocytes following exposure to mitogens, antigens, allogeneic or autologous cells, or soluble factors. Each of these protocols can be used in conjunction with an accompanying protocol, which contains methods to determine cell proliferation by incorporation of [(3)H]thymidine into DNA by nonradioactive methods, e.g., reduction of tetrazolium salts (MTT or WST-1). These protocols provide an estimate of cell proliferation indirectly by measuring DNA synthesis, and cell metabolic activity in an entire cell population, but no data on individual cells is obtained. A protocol for CFSE labeling allows direct detection of single proliferating cells and facilitates the quantification of cell divisions by flow cytometry according to the respective CFSE-dilution, and following costaining with fluorescent labeled antibodies, the characterization of subpopulations in the cell culture.

Measurement of proliferative responses of cultured lymphocytes.

Measurement of proliferative responses of human lymphocytes is a fundamental technique for the assessment of their biological responses to various stimuli. Most simply, this involves measurement of the number of cells present in a culture before and after the addition of a stimulating agent. This unit contains several different prototype protocols to measure the proliferative response of lymphocytes following exposure to mitogens, antigens, allogeneic or autologous cells, or soluble factors. Each of these protocols can be used in conjunction with an accompanying support protocol which contains methods for pulsing cultures with [3H]thymidine and determining incorporation of [3H]thymidine into DNA or assessing cell proliferation by nonradioactive methods, e.g., reduction of tetrazolium salts (MTT). The protocols described here provide an estimate of DNA synthesis and cell proliferation in an entire cell population, but do not provide information on the proliferation of individual cells. A protocol for CFSE labeling allows specific subpopulations of cells to be separated viably for further analysis.

Primary sclerosing cholangitis: summary of a workshop.

Primary sclerosing cholangitis (PSC) is a rare but important liver disease that leads to cirrhosis and need for liver transplantation in a high proportion of cases. The disease occurs in approximately 1 per 100,000 population per year, usually presents in adulthood, and affects men more often than women. Typical serum biochemical results, autoantibodies and liver biopsy are suggestive but not diagnostic of PSC, the diagnosis requiring cholangiographic demonstration of stricturing and dilatation of the intra- and/or extra-hepatic bile ducts. The natural history of PSC is variable, the average survival being 12 to 17 years. The cause of PSC is still unknown. Although considered an autoimmune disease, PSC has several atypical features and a strong genetic component. The therapy of PSC is unsatisfactory. Standard doses of ursodeoxycholic acid (UDCA) lead to improvements in biochemical abnormalities but not in histology, cholangiographic appearance or survival. Several innovative therapies have been tried in PSC, but with scant evidence of benefit. For patients with high grade strictures, endoscopic dilatation is beneficial. Liver transplantation is successful for end-stage liver disease due to PSC and improves survival. PSC may recur after transplantation but is rarely progressive. The most dreaded complication of PSC is cholangiocarcinoma. Diagnosis of this highly malignant tumor is difficult, and there are no biomarkers for its early detection. Liver transplantation for cholangiocarcinoma has an exceedingly poor outcome, although transplantation with neoadjuvant chemoirradiation holds promise in selected patients. Thus, significant opportunities remain for basic and clinical research into the cause, natural history, and therapy of PSC.

Prototypic disorders of gastrointestinal mucosal immune function: Celiac disease and Crohn's disease.

Celiac disease is a disorder of the small intestine characterized by chronic inflammation of the mucosa and protean clinical manifestations caused by loss of tolerance to dietary antigens. Two strongly associated cofactors have been identified: the presence of HLA-DQ2 or HLA-DQ8 in the host and specific antigenic peptides in the diet that are present in wheat, rye, and barley. Most patients have complete remission after dietary elimination of these foods. Crohn's disease is characterized by chronic, relapsing, recurrent, focal, transmural inflammation of the gastrointestinal tract that can lead to multiple serious problems requiring chronic medical and surgical therapy. Crohn's disease is associated with multiple genetic mutations, at least one of which has been clearly implicated in innate immunity. Multiple lines of evidence suggest that the disease involves abnormal immune responses to gut microbial flora.

Cutting edge: cyclooxygenase-2 activation suppresses Th1 polarization in response to Helicobacter pylori.

Helicobacter pylori infection causes a Th1-driven mucosal immune response. Cyclooxygenase (COX)-2 is up-regulated in lamina propria mononuclear cells in H. pylori gastritis. Because COX-2 can modulate Th1/Th2 balance, we determined whether H. pylori activates COX-2 in human PBMCs, and the effect on cytokine and proliferative responses. There was significant up-regulation of COX-2 mRNA and PGE(2) release in response to H. pylori preparations. Addition of COX-2 inhibitors or an anti-PGE(2) Ab resulted in a marked increase in H. pylori-stimulated IL-12 and IFN-gamma production, and a decrease in IL-10 levels. Addition of PGE(2) or cAMP, the second messenger activated by PGE(2), had the opposite effect. Similarly, stimulated cell proliferation was increased by COX-2 inhibitors or anti-PGE(2) Ab, and was decreased by PGE(2). Our findings indicate that COX-2 has an immunosuppressive role in H. pylori gastritis, which may protect the mucosa from severe injury, but may also contribute to the persistence of the infection.

19. Immunologic, gastroenterologic, and hepatobiliary disorders.

A number of gastrointestinal and hepatobiliary disorders are thought to have an immunologic basis; however, the level of understanding of immunopathogenesis varies widely in these disparate conditions. For some, such as celiac disease, both important genetic and environmental determinants have been identified as well as a specific treatment. For others, such as pernicious anemia and inflammatory bowel disease, animal models have provided important research advances. Two conditions, inflammatory bowel disease and sclerosing cholangitis, are strongly associated. Characteristic immunologic features of these diseases are critical for diagnosis. Although the majority of conditions are currently treated with nonselective anti-inflammatory and immunosuppressive agents, recent research has introduced novel biological agents, exemplified by the successful use of infliximab for Crohn's disease.