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M Möhlig

Publications and source records attributed to M Möhlig.

34 records · Page 2Linked to original sources
PubMed2001

Retinal photocoagulation does not influence intraocular levels of IGF-I, IGF-II and IGF-BP3 in proliferative diabetic retinopathy-evidence for combined treatment of PDR with somatostatin analogues and retinal photocoagulation?

Retinal photocoagulation reduces the incidence of severe visual loss in proliferative diabetic retinopathy (PDR). Reduced levels of VEGF/VPF might result in an improved function of the blood-retina barrier and cause a decrease of blood derived intraocular growth factors such as IGF-I. This study investigates whether retinal photocoagulation is able to normalize the concentrations of IGF-I, IGF-II and IGF-BP3 in the vitreous humor of patients undergoing vitrectomy. Levels of IGFs and the permeability marker, albumin, were measured in serum and vitreous of 52 patients. Three groups were compared: controls without proliferating eye disease (n = 19) and patients with PDR with (PDR+; n = 25) and without (PDR-; n = 8) previous retinal photocoagulation. IGF-I, IGF-II, IGF-BP3 and albumin were determined by immunological methods and were confirmed to be increased in patients with PDR compared to controls. Retinal photocoagulation influenced neither the intraocular concentration of the permeability marker albumin (PDR+: 253.2 +/- 46 mg/dl; PDR-: 256.4 +/- 66.5 mg/dl) nor the levels of IGFs (PDR+: IGF-I: 1.2 +/- 0.1 ng/ml; p = 0.38; IGF-II: 34.8 +/- 2.2 ng/ml; p = 0.1; IGF-BP3: 75.7 +/- 9.7 ng/ml; p = 0.27; PDR-: IGF-I: 1.1 +/- 0.2ng/ml; IGF-II: 29.3 +/- 5.2 ng/ml; IGF-BP3: 61.5 +/- 18.3 ng/ml). Systemic levels of albumin and IGFs were not changed significantly by retinal photocoagulation. These results demonstrate that previous retinal photocoagulation in patients undergoing vitrectomy does not functionally reestablish the blood-retina barrier despite decreases in VEGF/VPF. The lack of influence on intraocular concentrations of the serum-derived growth factors, IGF-I, IGF-II and IGF-BP3, might in part explain the failure of previous photocoagulation in the investigated patients. These results suggest that a combined treatment with retinal photocoagulation and growth hormone-lowering drugs, such as somatostatin analogues, could be a useful treatment, which may prevent further loss of visual acuity in patients with PDR.

Adult↗
PubMed2001

Loss of the antiangiogenic pigment epithelium-derived factor in patients with angiogenic eye disease.

Retinal neovascularization characterizes proliferative diabetic retinopathy (PDR). Pigment epithelium-derived factor (PEDF) has been shown to be a major antiangiogenic growth factor in the mammalian eye. PEDF expression is suppressed by hypoxia, and changes in PEDF have been correlated to the development of retinal neovascularization in animal models of hypoxic eye disease. However, whether this concept of a reduced angiogenesis inhibitor holds true in humans is as yet unclear. In this study, we analyzed the in vivo regulation of PEDF in patients with and without hypoxic eye disease. We used immunoblots to measure PEDF in ocular fluids obtained from 64 nondiabetic and diabetic patients. In addition, immunohistochemistry of PEDF was carried out in specimens of normal human retinas and retinas with various degrees of diabetic retinopathy. The PEDF concentrations in patients with PDR (P < 0.001) or extensive nondiabetic retinal neovascularization caused by retinal-vein occlusion (P < 0.001) were lower than in control patients. Levels of PEDF were replenished in PDR patients with previous retinal scatter photocoagulation compared with PDR patients without previous photocoagulation (P = 0.01). Immunohistochemistry revealed an interstitial staining pattern as expected for a secreted protein, with an intense staining in retinas of patients without proliferative eye disease. However, in patients with PDR, little or no staining was detectable. Our data strongly support the concept that retinal angiogenesis is induced by loss of the major angiogenesis inhibitor in the eye, PEDF, in combination with an increased expression of angiogenic growth factors such as vascular endothelial growth factor. Our findings suggest that substitution of angiogenesis inhibitors may be an effective approach in the treatment of PDR.

Aged↗
PubMed2000

Effect of estradiol on insulin secreting INS-1 cells overexpressing estrogen receptors.

BACKGROUND: Estrogen has been shown to have profound effects on insulin and glucose metabolism in vivo. Indeed, estrogens were recently shown to modulate ion channel and secretory activities in endocrine cells. DESIGN AND METHODS: To investigate whether estrogenic influences are caused by direct effects on pancreatic beta-cells, we equipped INS-1 insulinoma cells with estrogen receptors and monitored insulin content and Ca(2+) fluxes as well as basal and stimulated insulin secretion upon different stimuli including glucose, the Ca(2+) ionophore ionomycin, the Ca(2+) channel agonist BayK8644, the protein kinase C activator TPA, and the adenylate cyclase activator forskolin. RESULTS AND CONCLUSION: Our data reveal that estradiol has no significant direct effect on proliferation rate, insulin content, basal and stimulated insulin output as well as Ca(2+) fluxes of insulin secreting cells in vitro, indicating that in vivo responses to estrogen on insulin and glucose metabolism result from indirect betacytotropic effects.

Calcium↗
PubMed1999

Insulin resistance and impaired insulin secretion due to phosphofructo-1-kinase-deficiency in humans.

UNLABELLED: The etiology of non-insulin-dependent diabetes mellitus (NIDDM) is usually explained as a combination of peripheral insulin resistance and impaired beta-cell function. Phosphofructo-1-kinase (PFK1) is a rate limiting enzyme in glycolysis, and its muscle subtype (PFK1-M) deficiency leads to an autosomal recessively inherited disorder known as glycogenosis type VII or Tarui's disease. It was evaluated whether PFK1-M deficiency leads to NIDDM in humans. A core family of four was evaluated for PFK1-M deficiency by DNA- and enzyme-activity-analyses. All members underwent oral and intravenous glucose tolerance test (oGTT/ivgtt), as well as an insulin sensitivity test (IST) using octreotide. RESULTS: Father (46 years, BMI 22.4 kg/m2) and older son (19 years, BMI 17.8 kg/m5) showed homozygous PFK1-M deficiency, while mother (47 years, BMI 28.4 kg/m5) and younger son (13 years, BMI 16.5 kg/m5) were shown to be heterozygously PFK1-M-deficient on enzyme activity levels. DNA analysis revealed an exon 5-missense-mutation at one allele of all four members, and an exon 22-frameshift-mutation at the other allele of the two homozygously affected individuals. By oGTT the father showed impaired glucose tolerance, and the mother clinical diabetes. By ivGTT both parents and the older son had a decreased first phase insulin secretion, and a diminished glucose disappearance rate. The IST showed marked insulin resistance in both parents and the older son, and moderate resistance in the younger son, previously not described. CONCLUSION: PFK1-M-deficiency leads to a metabolic state typical for early NIDDM in homozygously affected humans, especially concerning insulin resistance and loss of first phase beta-cell insulin secretion, and may contribute to the manifestation of NIDDM in a subgroup of patients.

Adult↗
PubMed1999

Ca2+/calmodulin-dependent protein kinase II isoenzymes gamma and delta are both present in H+/K+-ATPase-containing rabbit gastric tubulovesicles.

Ca2+/calmodulin-dependent protein kinase II is thought to participate in M3 muscarinic receptor-mediated acid secretion in gastric parietal cells. During acid secretion tubulovesicles carrying H+/K+-ATPase fuse with the apical membrane. We localized Ca2+/calmodulin-dependent protein kinase II from highly purified rabbit gastric tubulovesicles using Ca2+/calmodulin-dependent protein kinase II isoform-specific antibodies, in vitro phosphorylation and pharmacological inhibition of Ca2+/calmodulin-dependent protein kinase II activity by the potent Ca2+/calmodulin-dependent protein kinase II inhibitor KN-62. The presence of Ca2+/calmodulin-dependent protein kinase II in tubulovesicles was shown by immunoblot detection of both Ca2+/calmodulin-dependent protein kinase II-gamma (54 kDa) and Ca2+/calmodulin-dependent protein kinase II-delta (56.5 kDa). The immunoprecipitated Ca2+/calmodulin-dependent protein kinase II from tubulovesicles showed Ca2+/calmodulin-dependent protein kinase activity by phosphorylating autocamtide-II, a specific synthetic Ca2+/calmodulin-dependent protein kinase II substrate. KN-62 inhibited the in vitro autophosphorylation of tubulovesicle-associated Ca2+/calmodulin-dependent protein kinase II (IC50 = 11 nM). During the search for potential Ca2+/calmodulin-dependent protein kinase II substrates we identified different proteins associated with tubulovesicles, such as synaptophysin and beta-tubulin immunoreactivity, which were identified using specific antibodies. These targets are known to participate in intracellular membrane traffic. Ca2+/calmodulin-dependent protein kinase II is thought to play an important role in regulating tubulovesicular motor activity and therefore in acid secretion.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1998

Purification and characterization of a Ca2+/calmodulin-dependent protein kinase II from hog gastric mucosa using a protein-protein affinity chromatographic technique.

A peripheral member of the Ca2+/calmodulin-dependent protein kinase II (CaMkinase II) group has been purified from hog gastric mucosa with the use of a novel affinity-chromatographic step. For the well known neural isotypes of CaMkinase II, it is proposed that the subunits form holoenzymes through a specific domain at the C-terminus called the 'association domain'. We immobilized a bacterially expressed association domain from CaMkinase II-delta-2 and used it as an affinity column. This matrix was used as the last step in a sequential enzyme purification procedure from hog gastric mucosa and yielded a homogeneous CaMkinase II which showed the typical physical and enzymatic properties of CaMkinase II. The enzyme activity showed a dependence on Ca2+ and calmodulin (apparent K0.5 = 2.7 microM and K0.5 = 0.02 microM, respectively). We found a subunit molecular mass of 61 kDa. An apparent native molecular mass of 310 kDa was calculated. The Stokes radius and the sedimentation coefficient were 6.7 nm and 11.2 S, respectively. Moreover, the isolated CaMkinase II was very well inhibited by the CaMkinase-II-specific inhibitors KN-62 (Ki = 0.52 nM) and KT5926 (Ki = 0.79 nM). The phosphorylation of several substrates revealed its multifunctionality. The purified CaMkinase II had an apparent Km for Mg-ATP of 24.0 microM and for autocamtide-II of 0.62 microM. CaMkinase II is considered as a strong candidate for regulating vesicle released quanta such as acid, neurotransmitters or insulin.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1997

Deficiency of phosphofructo-1-kinase/muscle subtype in humans impairs insulin secretion and causes insulin resistance.

Non-insulin-dependent diabetes mellitus (NIDDM) is caused by peripheral insulin resistance and impaired beta cell function. Phosphofructo-1-kinase (PFK1) is a rate-limiting enzyme in glycolysis, and its muscle subtype (PFK1-M) deficiency leads to the autosomal recessively inherited glycogenosis type VII Tarui's disease. It was evaluated whether PFK1-M deficiency leads to alterations in insulin action or secretion in humans. A core family of four members was evaluated for PFK1-M deficiency by DNA and enzyme-activity analyses. All members underwent oral and intravenous glucose tolerance tests (oGTT and ivGTT) and an insulin-sensitivity test (IST) using octreotide. Enzyme activity determinations in red blood cells showed that the father (46 yr, body mass index [BMI] 22. 4 kg/m2) and older son (19 yr, BMI 17.8 kg/m2) had a homozygous, while the mother (47 yr, BMI 28.4 kg/m2) and younger son (13 yr, BMI 16.5 kg/m2) had a heterozygous PFK1-M deficiency. DNA analyses revealed an exon 5 missense mutation causing missplicing of one allele in all four family members, and an exon 22 frameshift mutation of the other allele of the two homozygously affected individuals. The father showed impaired glucose tolerance, and the mother showed NIDDM. By ivGTT, both parents and the older son had decreased first-phase insulin secretion and a diminished glucose disappearance rate. The IST showed marked insulin resistance in both parents and the older, homozygous son, and moderate resistance in the younger son. PFK1-M deficiency causes impaired insulin secretion in response to glucose, demonstrating its participation in islet glucose metabolism, and peripheral insulin resistance. These combined metabolic sequelae of PFK-1 deficiency identify it as a candidate gene predisposing to NIDDM.

Adult↗
PubMed1997

Circulating tumor necrosis factor alpha is elevated in male but not in female patients with type II diabetes mellitus.

The cytokine tumor necrosis factor alpha (TNF alpha) was proposed to mediate obesity related insulin resistance upon production in fat cells and to participate in tissue remodelling leading to vascular complications upon being released by macrophages. To assess its putative role in diabetes we determined plasma levels of TNF alpha in 105 adult humans. Male nondiabetic subjects had significantly lower TNF alpha levels than female controls (4.4 +/- 0.3, n = 17 vs. 6.6 +/- 1.0 pg/ml, n = 13; p = 0.049). Men with NIDDM had elevated TNF alpha (6.7 +/- 0.6 pg/ml, n = 34) compared to nondiabetic subjects (4.4 +/- 0.3 pg/ml, n = 17; p = 0.012). Such a difference was not apparent in women. Levels of TNF alpha were correlated with serum triglyceride levels in male controls (r2 = 0.64; p = 0.007) but not in NIDDM. Neither body mass index nor glycosylated hemoglobin correlated with TNF alpha in any of the groups. The presence of retinopathy (p = 0.046) but not of neuropathy or nephropathy or macroangiopathy was associated with significantly elevated plasma TNF alpha. We conclude that plasma levels of TNF alpha are sex-dependent and that increased TNF alpha occurs in male but not female NIDDM and may participate in the development of diabetic complications.

Aged↗
PubMed1997

Insulinoma cells contain an isoform of Ca2+/calmodulin-dependent protein kinase II delta associated with insulin secretion vesicles.

The Ca2+/calmodulin dependent protein kinase II (CaM kinase II) is thought to play an important part in glucose-stimulated insulin secretion. To determine which of the known subtypes (alpha, beta, gamma, delta) occur in insulin-secreting cells, we amplified all types of CaM kinase II by RT-PCR and found the beta3-, gamma-, delta2- and delta6-subtypes in RINm5F insulinoma cells. None of the other 8 delta-subtypes was present. Antibodies generated against the bacterially expressed association domain of the delta2-subtype recognized the recombinant gamma and delta-subtypes. In INS-1 and RINm5F cells, as well as freshly isolated rat islets, only a 55-kDa protein corresponding in size to the delta2-subtype expressed in NIH3T3 fibroblasts was detected. The delta2-subtype therefore appears to represent the predominant subtype of CaM kinase II present in insulin secreting cells. The enzyme was primarily associated with cytoskeletal structures, and very little was present in the soluble compartment or detergent soluble fraction in INS-1- or RINm5F-cells. An analysis of its subcellular distribution was performed by sucrose and Nycodenz density gradient fractionation of INS-1 cells and detection of CaM kinase II delta by immune blots. The enzyme codistributed with insulin used as a marker for secretory granules but not with the lighter synaptic-like microvesicles detected with an antibody against synaptophysin, plasma membranes (syntaxin 1), lysosomes (arylsulfatase), or mitochondria (cytochrome c oxidase). CaM kinase II delta2 thus is identified as the subtype associated with insulin secretory granules and is likely to be involved in insulin secretion.

Animals↗
PubMed1995

Overexpression of calcium/calmodulin-dependent protein kinase II in insulinoma cells by use of a retroviral vector.

The manner in which increasing intracellular calcium levels lead to insulin exocytosis is not known. Possibly the signal is mediated by activation of calcium/calmodulin-dependent protein kinase II (CaM Kin II). In this work the establishment of an insulinoma cell line stably overexpressing CaM Kinase II subtype delta 2 by an ecotropic retroviral transduction system is described.

Calcium-Calmodulin-Dependent Protein Kinase Type 2↗
PubMed1994

Characterization of gamma- and delta-subunits of Ca2+/calmodulin-dependent protein kinase II in rat gastric mucosal cell populations.

We searched for the occurrence of a Ca2+/calmodulin-dependent protein kinase in rat gastric cell types as a likely member in the chain of gastrin- and muscarinic-receptor-mediated signal transmission. A Ca(2+)- and calmodulin-dependent phosphorylation of major 50, 60 and 100 kDa substrates was observed in parietal cell cytosol and a major 60 and 61 kDa protein doublet was found to bind 125I-calmodulin in 125I-calmodulin-gel overlays. A specific substrate of the multifunctional Ca2+/calmodulin-dependent protein kinase II, autocamtide II, was phosphorylated in a calmodulin-dependent manner. The specific inhibitor of this enzyme, KN-62, antagonized protein kinase activity. RNA extracted from gastric mucosal cells was shown to contain sequences of the gamma- and delta- but not alpha- and beta-subunits of the calmodulin-dependent kinase II, and mRNA of both subtypes was demonstrated in highly purified parietal, chief and mucous cells. A calmodulin-dependent kinase II composed of gamma- and delta-subunits is a likely mediator of Ca(2+)-dependent signal transmission in these populations of gastric cells.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1993

New isoforms of multifunctional calcium/calmodulin-dependent protein kinase II.

Calcium/calmodulin dependent protein kinase II (CaM kinase II) seems to act as an important regulator of intracellular signal transmission. Four subtypes, termed alpha to delta, have been cloned; some of them can exist as different splicing variants. All these isoforms share a great overall homology, and they contain 3 areas of low homology. We have identified 5 new variants of subtype delta so that the total number of different isoforms now adds up to 12. These variants are probably a result of different splicing and show several deletions in regard to subtype delta. The deletion sites do exactly match regions of low homology between the subtypes. This suggests a functional division of the CaM kinase II molecule into homologous and variable domains. The homologous domains are highly conserved. Therefore, it might be the case that the constitution of the variable domains is more significant for a certain isoform than its belonging to one of the 4 subtypes alpha to delta.

Alternative Splicing↗
PubMedDate not supplied

Novel and uncommon isoforms of the calcium sensing enzyme calcium/calmodulin dependent protein kinase II in heart tissue.

Calcium/calmodulin-dependent protein kinase type II (CaM kinase II) is an intracellular enzyme discovered several years ago in the brain which is obviously involved in intracellular signal transmission. Meanwhile it was detected in virtually every mammalian tissue. Several isoforms have been found to exist. Most recently the tissue distribution and the molecular structure of these isoforms have suggested that each form entails a particular function. In the present study we describe the identification, cloning, and nucleotide sequencing of two novel CaM kinase II isoforms which we discovered in rat heart. The presence of these additional subtypes makes the heart the organ which possesses the greatest number of different and unusual CaM kinase II isoforms throughout the body except for the brain. The importance of this finding is underscored by the fact that calcium is involved in the regulation of many crucial cardial parameters. The deduced amino acid sequence that we have obtained from the new CaM kinase II isoforms indicates a molecular organization which could make the design of subtype-specific inhibitory drugs for CaM kinase II possible. Such compounds would act similarly to but much more selectively than digitalis glycosides and would be likely to possess less side effects.

Amino Acid Sequence↗
PubMedDate not supplied

Insulin decreases human adiponectin plasma levels.

Insulin resistance and hyperinsulinemia are known atherosclerosis risk factors. The association between adiponectin plasma levels and obesity, insulinemia, and atherosclerosis has been shown. Thus, adiponectin may be a link between hyperinsulinemia and vascular disease. In vitro data demonstrated a reduction of adiponectin expression by insulin. However, it is still unclear whether insulin regulates adiponectinemia in vivo in humans. Five healthy male volunteers were studied. Circulating adiponectin levels were determined before and during hyperinsulinemic euglycemic clamp. Adiponectin was measured by radioimmunoassay. Hyperinsulinemia (85.0 +/- 33.2 at baseline vs. 482.8 +/- 64.4 pmol/l during steady state; p < 0.01) was achieved using a euglycemic hyperinsulinemic clamp, keeping blood glucose levels basically unchanged during the intervention (4.6 +/- 0.14 vs. 4.37 +/- 0.15 mmol/l, respectively; ns). We found a significant decrease of adiponectin plasma levels during the steady state of hyperinsulinemic euglycemic clamp (26.7 +/- 3.5 micro g/ml) compared to baseline levels (30.4 +/- 5 micro g/ml; p < 0.05). Hyperinsulinemia caused a significant decrease of adiponectin plasma levels under euglycemic conditions. Considering existing data about adiponectin dependent effects, hypoadiponectinemia might at least partly be a link between hyperinsulinemia and vascular disease in metabolic syndrome.

Adiponectin↗