Killer immunoglobulin receptor genes and their HLA-C ligand are associated with Type 1 diabetes in an Eastern Indian population.

AIM: Killer immunoglobulin-like receptors (KIRs) and their interaction with HLA class I ligands have been shown to be associated with Type 1 diabetes mellitus. The aim of our study was to investigate the influence of KIR genes and their HLA-C ligands for susceptibility to Type 1 diabetes in patients from Eastern India. METHODS: A total of 135 patients with Type 1 diabetes and 98 healthy subjects from Eastern India were typed for KIR genes and HLA-C ligands using PCR-based genotyping. The frequencies of these genes were compared between patients and controls. RESULTS: Comparison of KIR genes between Type 1 diabetes patients and healthy subjects revealed significantly different frequencies of KIRs 2DL2 and 2DS4. The presence of HLA-C1 was negatively associated with disease. The presence of both HLA-C1 and -C2 showed a negative association with Type 1 diabetes, whereas the absence of C1 and presence of C2 was positively associated with disease. Stratification analysis of HLA-C ligands and KIRs showed significant associations between Type 1 diabetes and 2DL2+/C1-, 2DL2-/C1+, 2DL3+/C1+, 2DL3+/C1- and 2DS2+/C1-. CONCLUSIONS: Our results suggest that the interaction of KIRs with HLA-C ligands are significant and certain combinations contribute to susceptibility to and protection against Type 1 diabetes.

Equivalency of tricyclic antidepressants in open-label neuropathic pain study.

OBJECTIVES: To compare adverse effects, tolerability and efficacy of the tricyclic antidepressants (TCAs) amitriptyline and nortriptyline in management of neuropathic pain due to peripheral neuropathy (PN). MATERIALS & METHODS: We performed a prospective open-label flexible-dosing comparison of monotherapy or adjuvant therapy using amitriptyline or nortriptyline in PN-associated neuropathic pain. Primary outcomes were quantitative adverse effects and discontinuation rates. Secondary outcomes assessed changes in pain severity, quality of life, disability, sleep efficacy, mood and anxiety, and global improvement. Assessments occurred at 3 and 6 months after initiation. Our hypothesis was that nortriptyline would have better tolerance than amitriptyline. RESULTS: A total of 228 PN patients were enrolled approximately equally for monotherapy and adjuvant therapy. Adverse effects and discontinuation rates were similar between amitriptyline and nortriptyline interventions. Weight gain was more common with amitriptyline, while nortriptyline use was associated with greater prevalence of dry mouth. Secondary outcome measures were similar in both groups, demonstrating improvement from baseline. CONCLUSIONS: Amitriptyline and nortriptyline are equivalent for overall adverse effects and discontinuation rates. Either TCA should be equally considered for use in neuropathic pain due to PN. When used as monotherapy or as part of adjuvant therapy, either TCA can be expected to provide approximately 23-26% visual analog scale pain reduction if tolerated. Discontinuations due to inefficacy or adverse effects can be anticipated in 26-37% of patients initiated on either TCA for PN-associated neuropathic pain.

Caspase-3 deficiency reveals a physiologic role for Smac/DIABLO in regulating programmed cell death.

Inhibitor of apoptosis protein (IAP)-binding proteins such as Grim, Reaper and HID have been shown to exert a critical role in regulating caspase activity in species such as D. Melanogaster. However, a comparable role for the mammalian homologue of second mitochondrial-derived activator of caspase/direct IAP-binding protein with low pI (Smac/DIABLO) has yet to be clearly established in vivo. Despite tremendous interest in recent years in the use of so-called Smac mimetics to enhance chemotherapeutic potency, our understanding of the true physiologic nature of Smac/DIABLO in regulating programmed cell death (PCD) remains elusive. In order to critically evaluate the role of Smac/DIABLO in regulating mammalian PCD, deficiency of caspase-3 was used as a sensitizing mutation in order to reduce aggregate levels of executioner caspase activity. We observe that combinatorial deletion of Diablo and Casp3, but neither alone, results in perinatal lethality in mice. Consistent with this, examination of both intrinsic and extrinsic forms of PCD in lines of murine embryonic fibroblasts demonstrate that loss of Smac/DIABLO alters both caspase-dependent and caspase-independent intrinsic PCD. Comparative small interfering RNA inhibition studies of X-linked inhibitor of apoptosis, cellular inhibitor of apoptosis (cIAP)-1, cIAP-2, caspase-6 and -7 in both wild-type and Casp3/Diablo DKO mouse embryonic fibroblast lineages, supports a model in which Smac/DIABLO acts to enhance the early phase executioner caspase activity through the modulation of inhibitory interactions between specific IAP family members and executioner caspases-3 and -7.

Excitatory tonus is required for the survival of granule cell precursors during postnatal development within the cerebellum.

In addition to protective effects within the adult central nervous system (CNS), in vivo application of N-methyl-d-aspartate inhibitors such as (+) MK-801 have been shown to induce neurodegeneration in neonatal rats over a specific developmental period. We have systematically mapped the nature and extent of MK-801-induced neurodegeneration throughout the neonatal murine brain in order to genetically dissect the mechanism of these effects. Highest levels of MK-801-induced neurodegeneration are seen in the cerebellar external germinal layer; while mature neurons of the internal granule layer are unaffected by MK-801 treatment. Examination of external germinal layer neurons by electron microscopy, terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling (TUNEL) and bromodeoxyuridine (BrdU) labeling, and caspase-3 activation demonstrate that these neurons die through the process of programmed cell death soon after they exit from the cell cycle. Significantly, ablation of caspase-3 activity completely inhibited the MK-801-induced (and developmental) programmed cell death of external germinal layer neurons. Similar to caspase-3, inactivation of muscarinic acetylcholine receptors in vivo using scopolamine inhibited MK-801-induced programmed cell death. By contrast, the GABAergic agonist diazepam, either alone or in combination with MK-801, enhanced programmed cell death within external germinal layer neurons. These data demonstrate that, in vivo, cerebellar granule neurons undergo a dramatic change in intracellular signaling in response to molecules present in the local cellular milieu during their first 24 h following exit from the cell cycle.

No association of SUMO4 M55V with autoimmune diabetes in Asian-Indian patients.

Autoimmune diabetes [type 1 diabetes mellitus (T1DM), latent autoimmune diabetes in adults (LADA) and part of malnutrition-related diabetes] has been shown to have genetic predisposition. Studies in IDDM 5 have lead to the discovery of a novel polymorphism 163 A-->G, of SUMO4 (small ubiquitin-related modifier) gene, associated with risk to T1DM in Asians, but not in Caucasians. We studied patients with T1DM (n = 134), patients with LADA (n = 101), patients with malnutrition-modulated diabetes mellitus (n = 66) and patients with fibrocalculous pancreatic diabetes (n = 43) and healthy controls subjects (n = 114) from Cuttack, India. Polymerase chain reaction-sequence-specific primer (PCR-SSP) was used to amplify the 163 A-->G sequences. Restriction fragment length polymorphism (RFLP) was performed using restriction enzyme Taq I (PCR-RFLP). Differences in the allelic frequencies of the A and the G alleles were tested statistically using Fisher's exact test or chi-squared test wherever appropriate. P-values were considered significant when equal to or less than 0.05. No significant association was detected between SUMO4 M55V and T1DM susceptibility in Asian-Indians. Comparison of the A and G alleles with HLA DR3-DR4 did not result in any significant P-values. No significant association was found between SUMO4 M55V and LADA or malnutrition-related diabetes mellitus (MRDM). Our results show that Asian-Indians with T1DM are different from other Asian populations. Asian-Indians show more similarity to Caucasians with respect to the association of SUMO4 M55V variant in T1DM. Association studies on Asian-Indian patients with LADA and MRDM showed no significant difference in the presence of the A and the G alleles when compared to healthy controls.

Genetics of latent autoimmune diabetes in adults.

Slowly progressive insulin-dependent diabetes mellitus (IDDM), like classical IDDM, is also associated with genetic markers. HLA-DR3 but not DR4 is associated with latent autoimmune diabetes in adults (LADA). In GAD65 antibody-positive Finnish LADA patients, DQB1*0302 is positively associated with the disease. Alleles of the MHC class I chain-related A (MICA) gene located centromeric to the HLA-B gene is associated with LADA. Allele 5.1 of MICA was associated with both LADA and adult-onset Italian IDDM patients when compared to controls. This finding was also observed in Indian and Latvian patients with LADA. These findings suggest that certain genetic markers distinguish LADA better.

Molecular mechanisms involved in the etiopathogenesis of malnutrition-modulated diabetes mellitus.

MMDM patients are typically young at onset with low body mass index, require insulin treatment for glycemic control, have insulin resistance, and do not develop ketosis on withdrawal of insulin. WHO's revised classification in 1999, based on the etiopathogenesis of the disease, identifies only two categories: type 1 diabetes and type 2 diabetes. MMDM could be considered as type 1b diabetes. Genetic and immunological studies were done on MDDM patients (n = 72) from Cuttack and healthy controls to understand and to justify its inclusion in the category of type 1b diabetes. Antibodies (Abs) to tyrosine pyrophosphatase (IA2-Abs), glutamate decarboxylase 65 (GAD65-Abs), and other minor markers like ICA12 Abs and tissue transglutaminase Abs (TTG-Abs) were studied. HLA-DR and DQ were studied for the genetic markers. Of the MMDM patients 30% were positive for either GAD65 or IA-2 antibodies, and 14% were positive for ICA12 antibodies. All three antibody markers together accounted for 39% of PDDM patients, as some patients were positive for more than one autoantibody. TTG antibodies (specific for Celiac disease) were present in 14/71 (20%) of MMDM patients compared to 3/122 (2%) controls. All four autoantibodies accounted for 53% of PDDM patients, leaving 47% of patients free of known autoantibodies. The autoantibody-negative PDDM patients were analyzed for HLA and MICA markers, showing that DR7-DQ9 and MICA allele 9 are increased in this group compared to healthy controls, which suggests an autoimmune response to an unknown dietary autoantigen. We conclude from our data that an autoimmune mechanism is involved in the etiology of MMDM. In addition, the presence of silent celiac disease seen with MMDM patients, which has not yet been reported, is significant. It is important to note that subclinical celiac disease exists with diabetes mellitus and must be considered in the diagnosis of MMDM.

Clinicoepidemiological and biochemical profile of malnutrition-modulated diabetes mellitus.

Malnitrition-modulated diabetes mellitus ((MMDM) was previously known as protein-deficient diabetes mellitus (PDDM). Its clinical picture is similar to that of type 1 diabetes, but it develops over a background of chronic malnutrition from childhood. In spite of severe hyperglycemia, ketonuria never occurs. MMDM patients are extremely lean and require high doses of insulin-over 2.0 U/kg/day for good glycemic control. Even when optimally controlled, these patients maintain their leanness. Infections of the skin and soft tissues and pulmonary tuberculosis are often seen, whereas micro- and macrovascular complications are rare, even after long-term follow-up. Ultrasonographic evaluation of the abdomen clearly differentiates MMDM from fibrocalculous pancreatic diabetes. Absence of ketonuria and ketosis despite very severe hyperglycemia in emaciated young subjects is the most significant marker of MMDM.

Immunogenetic studies on malnutrition-modulated diabetes mellitus.

Genetic studies of malnutrition-related diabetes are few. We have analyzed the HLA class II gene polymorphism in malnutrition-modulated diabetes mellitus (MMDM), which was previously referred to as protein-deficient diabetes mellitus (PDDM) in the 1985 WHO classification. Insulin-dependent diabetes mellitus (IDDM) is a polygenic disorder with an autoimmune basis for disease development. In addition to HLA, a second susceptibility locus for IDDM has been identified to lie in the major histocompatibility class III region. Both IDDM and MMDM in eastern Indians are associated with DR3-DQ2 but not DR4-DQ8. The presence of autoantibodies to IDDM autoantigens in clinical MMDM either identifies the slow-onset form of IDDM or suggests autoimmunity different from that in IDDM. Our study demonstrates that the presence of GAD65 antibody and DR3-DQ2 positivity in MMDM patients identifies the underlying autoimmune mechanism in the etiology in eastern India. In autoantibody-negative MMDM patients an association with DR7-DQ2 is identified. The date obtained also indicate the possibility that MMDM can coexist with IDDM in these patients and that malnutrition could be one of the reasons for the slower onset in IDDM-prone individuals. The association of DR7-DQ2 suggests that there is a different immunogenetic background to MMDM than to IDDM. MICA is located in the MHC class I region and is expressed by monocytes, keratinocytes, and endothelial cells. Sequence determination of MICA gene identifies trinucleotide repeat (GCT) microsatellite polymorphism in exon 5. Five alleles with 4, 5, 6, and 9 repetitions of GCT or 5 repetitions of GCT with 1 additional nucleotide insertion (GGCT) are identified. The alleles are A4, A5, A5.1, A6, and A9. We studied the association of MICA alleles with IDDM (n = 52) and MMDM (n = 41) patients and healthy controls (n = 73) from Cuttack, eastern India. MICA was typed by PCR amplification, and fragment sizes were determined in an ABI prism DNA sequencer. Allele 9 of MICA is positively and allele 4 negatively associated with MMDM patients compared to controls. Allele 5 is positively associated with IDDM (OR 2.64, P < 0.05) when compared to controls. Our findings suggest that MMDM is immunogenetically different from IDDM in eastern India and that MIC-A is important in the pathogenesis of MMDM patients from Cuttack in eastern India.

Autoantibodies to tissue transglutaminase in patients from eastern India with malnutrition-modulated diabetes mellitus, insulin-dependent diabetes mellitus, and non-insulin-dependent diabetes mellitus.

Antibodies to tyrosine phosphatase (IA2-Ab) and glutamate decarboxylase 65 (GAD65-Ab) are major markers for IDDM in Caucasians. TTG-Ab is specific for celiac disease. Celiac disease is caused by ingestion of the protein gliadin, a component of wheat gluten, and usually resolves on its withdrawal. Ten to twenty percent of celiac disease patients also have IDDM. The aim of the study was to estimate the prevalence of TTG-Ab in MMDM (n = 71), IDDM (n = 74), and NIDDM (n = 216) and 122 controls from Cuttack in eastern India. MMDM patients are typically young at onset with low body mass index, require insulin for glycemic control, have insulin resistance, and do not develop ketosis on withdrawal of insulin. TTG-Ab was evaluated by radioimmunoassay using in vitro translated recombinant human 35S-TTG. In controls, TTG-Ab was present in 3/122 (2%); in MMDM, TTG-Ab was present in 14/71 (20%); 11/74 (15%) IDDM (P < 0.05 vs. controls) and 23/216 (11%) NIDDM (P < 0.05 vs. controls) were also positive for TTG-Ab. We conclude that MMDM, IDDM, and NIDDM patients from Cuttack have a significantly high proportion of TTG-Ab compared to healthy controls. The highest significance is seen with MMDM patients. It is important to note that subclinical celiac disease must be considered in the differential diagnosis of MMDM.

MHC class I chain-related gene a alleles distinguish malnutrition-modulated diabetes, insulin-dependent diabetes, and non-insulin- dependent diabetes mellitus patients from eastern India.

Insulin-dependent diabetes mellitus (IDDM) is a polygenic disorder with an autoimmune basis for disease development. In addition to HLA, a second susceptibility locus for IDDM has been identified to lie in the major histocompatibility class III region. MIC-A is located in the MHC class III region and is expressed by monocytes, keratinocytes, and endothelial cells. Sequence determination of the MIC-A gene identifies trinucleotide repeat (GCT) microsatellite polymorphism in exon 5. Five alleles with 4, 5, 6, and 9 repetitions of GCT or 5 repetitions of GCT with 1 additional nucleotide insertion (GGCT) are identified. The alleles are A4, A5, A5.1, A6, and A9. The aim of our study was to find the association of MIC-A alleles with IDDM, malnutrition-modulated diabetes mellitus (MMDM), and non-insulin-dependent diabetes mellitus (NIDDM) patients. IDDM (n = 52), MMDM (n = 41), NIDDM (n = 212), and healthy controls (n = 73) from Cuttack, in eastern India, were studied. Of the 212 NIDDM patients analyzed, 96 of them were found to be positive for either GAD65 or IA-2 antibodies. Autoantibodies to GAD65 and IA-2 were measured by radioligand binding assay using (35)S-labeled recombinant human GAD65 and IA-2 in an in vitro transcription/translation system. Autoantibody-positive NIDDM patients (n = 96) and adult healthy controls for NIDDM (n = 113) were also compared. These autoantibody-positive NIDDM patients are considered as slow-onset IDDM or latent autoimmune diabetes in adults (LADA) patients. The samples were analyzed for MIC-A by PCR amplification, and fragment sizes were determined in an ABI prism DNA sequencer. The results of the MIC-A typing are: allele 9 of MIC-A is positively associated (OR 3.62; P < 0.001), and allele 4 is negatively associated (OR 0.31; P < 0.05) with MMDM patients compared to controls. Allele 5 is positively associated with IDDM (OR 2.64; P < 0.05) when compared to controls. Allele 5.1 is positively associated in the autoantibody-positive NIDDM patients compared to adult controls. Our findings of a significant increase of allele A9 in MMDM patients compared to healthy controls suggest that MMDM is immunogenetically different from IDDM in eastern India. MIC-A is important in the pathogenesis of MMDM patients from Cuttack. MIC-A alleles distinguish acute-onset IDDM from slow-onset IDDM, indicating that this molecule may be important for delaying the onset of IDDM with the result that these patients are diagnosed clinically as NIDDM.

Peptide YY/neuropeptide Y Y1 receptor expression in the epithelium and mucosal nerves of the human colon.

OBJECTIVES: Peptide YY is an abundant distal gut hormone which regulates secretion, motility, and possibly epithelial proliferation in the gut. Though messenger RNA for the peptide YY Y1 receptor subtype occurs in the basal colonic crypts of humans, peptide YY receptors themselves have not been clearly localized within the adult human gastrointestinal tract. Using an antiserum directed against the C-terminus of the Y1 receptor we determined the actual extent of Y1 receptor protein expression in the human colon in order to identify areas targeted for peptide YY effects and suggest additional physiological roles for PYY in the human gut. RESULTS: Y1 receptor protein expression was seen throughout the colonic epithelium along its basolateral aspect. There was an unexpected dense distribution of Y1 receptor immunoreactivity in varicose fibers within the mucosa. Staining was also noted in nerve fibers of the muscularis mucosae, in the submucous and myenteric plexuses, and in nerves in the muscularis propria. CONCLUSIONS: Widespread distribution of Y1 receptors in the colonic epithelium and mucosal nerve fibers suggests diverse regulatory roles for peptide YY in modulating epithelial function as well as secretomotor reflexes in response to lumenal peptide YY-release signals.

Association of HLA class II alleles with different subgroups of diabetes mellitus in Eastern India identify different associations with IDDM and malnutrition-related diabetes.

Genetic studies of Malnutrition related diabetes are few. We have analyzed HLA class II gene polymorphism in different types of diabetes mellitus patients from Cuttack in Eastern India. Patients with insulin-dependent diabetes mellitus (IDDM), non-insulin-dependent diabetes mellitus (NIDDM) and malnutrition-related diabetes mellitus (MRDM), which is subdivided into protein-deficient diabetes mellitus (PDDM) and fibrocalculous pancreatic diabetes (FCPD), were studied and their associations with autoantibody markers. IDDM and PDDM were associated with DR3 and DQ2 but not DR4 and DQ8. FCPD was positively associated with DQ9 (A*0201-B*0303). The association of DQ9 with FCPD suggests differences in the genetic background for susceptibility between IDDM and MRDM in the Cuttack population. There is no association seen between HLA-DR-DQ and NIDDM patients from Eastern India. Clinical classification of diabetes into IDDM, NIDDM and MRDM does not identify the underlying pathological mechanisms. Presence of autoantibodies to IDDM autoantigens in clinical MRDM and NIDDM identifies the slow-onset form of IDDM. Due to the absence of autoantibody assays for diagnosis of IDDM in India, slow onset IDDM is not diagnosed and the patients are classified as NIDDM or MRDM. Our study demonstrates that the presence of GAD65 antibody and DR3-DQ2 positivity in MRDM and NIDDM patients in Eastern India would suggest the presence of slow-onset IDDM. Our data would indicate alternatively that MRDM can coexist with IDDM in these patients and malnutrition could be one of the reasons for the slower onset in IDDM-prone individuals.

Melanotic neuroectodermal tumor of the epididymis. A case report.

Melanotic neuroectodermal tumors occur very rarely in the epididymis, only eleven cases having been reported to date in the world literature. An additional case is reported which we believe is the first from India.