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D Engelhardt

Publications and source records attributed to D Engelhardt.

At least 19 recordsLinked to original sources
PubMed2000

Induction of oral tolerance in splenocyte recipients toward pretransplant antigens ameliorates chronic graft versus host disease in a murine model.

Chronic graft versus host disease (cGVHD) is a major complication that can develop after bone marrow transplantation. It involves an immune-mediated attack by transplanted donor lymphocytes, and often results in inflammatory damage of host target organs. Immune hyporesponsiveness induced by oral antigen administration has been recently shown to prevent the development of cGVHD in a murine model. The aim of this study was to evaluate whether tolerance induction in bone marrow transplant (BMT) recipients after transplantation, toward their pretransplant antigens, can alleviate preexisting cGVHD in a mouse model. cGVHD was generated by infusing 2.5 x 10(7) splenocytes from B10.D2 donor mice, to sublethally irradiated (6 Gy) BALB/c recipient mice, which differ by minor histocompatibility antigens. Transplantation resulted in cGVHD, with characteristic scleroderma-like cutaneous fibrosis, increased skin collagen content, decreased body weight, and hepatic and small bowel inflammation. Oral tolerance was induced by feeding recipient BALB/c mice with proteins extracted from BALB/c splenocytes for 11 days after B10.D2 splenocyte transplantation. Tolerance induction was evidenced by a significant reduction in mixed lymphocyte response of effector splenocytes from tolerant BALB/c mice transplanted with B10.D2 splenocytes against BALB/c target splenocytes. Oral tolerance decreased skin collagen deposits. Reduction of collagen alpha1(I) gene expression and skin collagen were shown by in situ hybridization and histochemistry, respectively. Liver and bowel biopsy specimens revealed less inflammation. Serum IL-10 levels were higher in tolerant mice than in controls, whereas IFNgamma was significantly reduced. Oral tolerance of BMT recipients toward their pretransplant antigens after splenocyte transplantation down-regulated the immune attack by transplanted cells, thus ameliorating cGVHD.

Administration, Oral↗
PubMed2000

Overexpression of insulin-like growth factor-binding protein-2 results in increased tumorigenic potential in Y-1 adrenocortical tumor cells.

Increased concentrations of insulin-like growth factor-binding protein-2 (IGFBP-2) have been observed in human malignancies including adrenocortical carcinomas. To elucidate the functional consequences of IGFBP-2 overexpression, we have stably transfected the cDNA of murine IGFBP-2 in mouse adrenocortical tumor cells (Y-1). Long-term overexpression of IGFBP-2 was associated with significant morphological alterations, enhanced cell proliferation, and increased cloning efficiency as compared with mock transfected control cells. The enhanced proliferation of IGFBP-2 secreting clones was independent of exogenous insulin-like growth factors (IGFs). These data suggest that elevated levels of IGFBP-2 may contribute to the highly malignant phenotype of adrenocortical cancer by a thus far unknown, presumably IGF-independent, mechanism.

Adrenal Cortex Neoplasms↗
PubMed2000

A long-term hepatitis B viremia model generated by transplanting nontumorigenic immortalized human hepatocytes in Rag-2-deficient mice.

Development of new therapies for human hepatitis B virus infection (HBV) would be greatly facilitated by the availability of a suitable small-animal model for HBV virus production in vivo. To develop a murine model for HBV production, we established an immortalized, cloned liver cell line by transferring the Simian Virus 40 Large T-Antigen into primary human hepatocytes. These cells were stably transfected with a full-length HBV genome to generate a clone that expresses HBV genes and replicates HBV. The HBV-producing cells were transplanted into the livers of mice with combined immunodeficiency (Rag-2 deficient) by intrasplenic injection. Survival of the engrafted human hepatocytes was shown in several ways: fluorescent in situ hybridization (FISH) with a human-chromosome-specific DNA probe (human alpha satellite), dot-blot hybridization of the genomic DNA extracted from liver biopsy specimens with a human-specific Alu repetitive DNA probe, Blur-8, as well as with an HBV DNA probe, and secretion of human proteins into plasma. Histological examination of mouse liver up to 8 months following human cell transplant shows completely normal architecture. Determination of plasma HBV DNA levels indicated that engrafted cells secreted 3x10(7) to 3x10(8) virions per mL into the blood, and HBsAg was detected in plasma. This new murine model of HBV viremia should be useful for in vivo HBV studies.

Animals↗
PubMed2000

Oral tolerization ameliorates liver disorders associated with chronic graft versus host disease in mice.

In chronic graft versus host disease (cGVHD), an immune attack by transplanted donor lymphocytes results in damage of host target organs. A disbalance between proinflammatory (Th1) and anti-inflammatory cytokines (Th2) plays an important role in the pathogenesis. Immune hyporesponsiveness induced by oral antigen administration has been shown to suppress autoimmunity. We evaluated the efficacy of oral tolerization in preventing cGVHD in a mouse model. cGVHD was generated by infusing 2.5 x 10(7) splenocytes from B10.D2 donor mice, to sublethally irradiated (6 Gy) BALB/c recipient mice, which differ in minor histocompatibility antigens. The transplantation resulted in cGVHD, with characteristic hepatic and small bowel inflammation, and increased skin collagen content and fibrosis. Oral tolerance was induced by feeding donor B10.D2 mice with proteins extracted from BALB/c splenocytes at 50 microg/d per mouse for 11 days before transplantation. Tolerization was evidenced by reduction in mixed lymphocyte response of effector splenocytes from tolerized B10.D2 mice against BALB/c target splenocytes. Liver and small bowel biopsy specimens revealed much less inflammation. Oral tolerization prevented weight and subcutaneous fat loss, reduced thickening, and skin collagen deposits. Reduction of collagen alpha1 (I) gene expression was shown by in situ hybridization. Serum interleukin 10 (IL-10) levels measured significantly higher in tolerized mice than in controls, whereas interferon gamma (IFN-gamma), IL-2, and tumor necrosis factor alpha (TNF-alpha) were reduced significantly. Oral tolerization of splenocyte donors towards recipient-strain splenocytes ameliorated cGVHD of the liver, small intestine, and skin. A cytokine shift from a proinflammatory to an anti-inflammatory pattern may play a role in down-regulation of the immune-mediated target organ damage.

Animals↗
PubMed2000

Interleukin-3 and interleukin-6 stimulate bovine adrenal cortisol secretion through different pathways.

Several interleukins have been reported to play a major role in the regulation of steroid secretion at all three levels of the hypothalamic-pituitary-adrenal axis. The objective of this study was to investigate the effect of interleukin-3 (IL-3) and interleukin-6 (IL-6) on cortisol secretion of bovine adrenocortical cells in primary culture under serum-free conditions. Both IL-3 and IL-6 stimulated basal cortisol secretion dose-dependently to a similar extent at a similar time course. After incubation with IL-3 or IL-6 at concentrations of 100 microg/l, a maximum 4.1-fold increase of the cortisol secretion was reached after 12 h (P<0.01). Coincubation of IL-3 and IL-6 (100 microg/l) revealed no significant synergism. To elucidate a possible involvement of arachidonic acid metabolites in the signal transduction, we coincubated IL-3 or IL-6 together with the cyclo-oxygenase inhibitor indometacin or the lipoxygenase inhibitor nordihydroguaiaretic acid (NDGA). Coincubation with indometacin completely abolished the stimulatory effect of IL-6 but had no effect on IL-3 stimulated cortisol secretion. In contrast, specific inhibition of the lipoxygenase system by nordihydroguaiaretic acid blocked IL-3 stimulated steroidogenesis while the effect of IL-6 was not affected. Neither IL-3 nor IL-6 altered cAMP levels significantly, whereas ACTH significantly induced cAMP levels in parallel to its steroidogenic effect. In conclusion, our data indicate that IL-3 and IL-6 stimulate the steroid secretion of bovine adrenocortical cells to a similar extent and with a similar time course. However, the effects of IL-3 and IL-6 are mediated through different, cAMP-independent pathways. While the stimulatory effect of IL-3 seems to be dependent on the lipoxygenase pathway, the effect of IL-6 on adrenocortical cortisol secretion is mediated through the cyclo-oxygenase pathway.

Adrenal Cortex↗
PubMed2000

Treatment of experimental colitis by oral tolerance induction: a central role for suppressor lymphocytes.

OBJECTIVE: Inflammatory bowel diseases (IBD) are immune-mediated disorders wherein an imbalance between proinflammatory (Th1) and antiinflammatory (Th2) cytokines is thought to play a role in the pathogenesis. The aim of this study was to test whether induction of oral tolerance to proteins extracted from inflammatory colon alleviates experimental colitis, and whether oral tolerization mediated by suppressor cells can induce immune tolerance. METHODS: Colitis was induced in rats by intracolonic instillation of trinitrobenzenesulfonic acid (TNBS). Rats received five oral doses of colonic proteins extracted from TNBS-colitis colonic wall. Splenocytes harvested from tolerized and control rats were transplanted into irradiated naive rats. RESULTS: Feeding of colitis-extracted proteins ameliorated colonic inflammation, as shown by reduction of colonic ulcerations, as well as decreased diarrhea, intestine and peritoneal adhesions, wall thickness, and edema. A marked reduction of the fraction of injured colonic area and colon weight, and decrease in colon weight, were observed in tolerized rats versus controls. Histological parameters for colitis were markedly improved in tolerized animals that showed significant reduction in inflammatory response and mucosal ulcerations. Tolerized rats developed an increase in TGFbeta1 and a decrease in IFNgamma serum levels. TNBS-induced colitis was significantly attenuated in naive recipients of splenocytes from tolerized rats, compared with rats that received splenocytes from control donors. CONCLUSIONS: Induction of oral tolerance to colitis-extracted proteins downregulates the anticolon immune response, thereby ameliorating experimental colitis. Suppressor lymphocytes mediate the tolerance by induction of a shift from a proinflammatory to an antiinflammatory immune response.

Administration, Oral↗
PubMed1999

Postnatal overexpression of insulin-like growth factor II in transgenic mice is associated with adrenocortical hyperplasia and enhanced steroidogenesis.

The influence of postnatal insulin-like growth factor II (IGF-II) overexpression on adrenal growth and function was investigated in 3-month-old male phosphoenolpyruvate carboxykinase (PEPCK) promoter human IGF-II transgenic mice, which are characterized by 4-to 6-fold elevated postnatal IGF-II serum levels. Plasma corticosterone levels of PEPCK-IGF-II transgenic mice were 2-fold higher than in age- and sex-matched controls, both in the morning (7.4 +/- 1.5 vs. 17.8 +/- 3.9 ng/ml, P < 0.01) and in the evening (33.3 +/- 6.5 vs. 65.3 +/- 12 ng/ml, P < 0.01). When PEPCK-IGF-II transgenic mice were subjected to an ACTH challenge, corticosterone levels were stimulated 6-fold, to 396 +/- 17 ng/ml after 60 min, compared with 230 +/- 24 ng/ml in the control group. In contrast to corticosterone, plasma ACTH levels were similar in transgenic and control mice, excluding an indirect effect of IGF-II at the hypothalamic or pituitary level. In vitro, the basal and ACTH-induced corticosterone production of adrenal glands from transgenic mice was higher (2-fold and 1.8-fold, respectively) than that of control organs. However, when normalized for adrenal weight, the in vitro corticosterone secretion was similar in both groups. At autopsy, adrenal weights of transgenic mice were significantly greater than those of control adrenal glands (3.3 +/- 0.2 vs. 2.0 +/- 0.2 mg, P < 0.01, n = 10). Furthermore, a local expression of human IGF-II could be demonstrated in transgenic adrenal glands by RT-PCR, whereas in normal adult mice, no adrenal expression of IGF-II was detected. Stereological investigation of adrenal glands from another set of PEPCK-IGF-II transgenic mice and controls (6-month-old males) demonstrated that the increase in adrenal weight in transgenic mice is mainly caused by a 50% increase in the number of zona fasciculata cells, whereas cell volume and zonation of transgenic adrenal glands remained unchanged. In conclusion, our data indicate that postnatal overexpression of IGF-II induces an increased adrenal weight and elevated corticosterone serum levels, presumably by a direct mitogenic effect of IGF-II on adrenocortical fasciculata cells.

Adrenal Glands↗
PubMed1999

Characterization of human insulin-like growth factor-binding proteins by two-dimensional polyacrylamide gel electrophoresis and Western ligand blot analysis.

The insulin-like growth factor (IGF)-binding proteins (IGFBPs) from adult human serum, amniotic fluid, and cerebrospinal fluid were analyzed by a modified two-dimensional gel electrophoresis followed by Western ligand blotting. The samples were subjected to immobilized pH gradient isoelectric focusing in the first dimension, followed by nondenaturing SDS-PAGE in the second dimension and autoradiography after ligand blotting with [125I]IGF-I or [125I]IGF-II. The identity of the binding proteins was confirmed by immunoblotting and immunoprecipitation with specific antibodies. Using this method, all six human high affinity IGFBPs could be clearly separated from each other according to their molecular mass and isoelectric points (pI). All IGFBPs exhibited a variety of specific pI isoforms, which presumably represent posttranslational modifications. In adult human serum, glycosylated IGFBP-3 is found as a broad band of spots with molecular masses of 41 and 45 kDa and a pI in the range of 4.8-8.2. The two IGFBP-3 bands could be reduced to a single 36-kDa band after deglycosylation (pI 6-9). Furthermore, the specific spots for IGFBP-2 (33 kDa; pI 6.2-7.1) and deglycosylated IGFBP-4 (24 kDa; pI 6.3, 6.5, and 6.8) were found with their expected molecular masses. Additionally, the diffuse bands around 30 kDa, found in one-dimensional Western ligand blotting, could be clearly separated into distinct groups of specific spots representing IGFBP-1 (30 kDa; pI 4.0-4.8), IGFBP-6 (30 kDa; pI 4.8-5.8), glycosylated IGFBP-4 (29 kDa; pI 6.1 and 6.3), and IGFBP-5 (30/31 kDa; pI 6.4-8). As expected, IGFBP-6 was visible only when IGF-II was used as radioligand. In conclusion, two-dimensional gel electrophoresis followed by Western ligand blotting allows identification of all six high affinity IGFBPs with their isoforms on the basis of their characteristic molecular masses and pI, especially in the range of 30 kDa. This technique can be rapidly performed with small amounts of complex biological fluids and is a powerful tool for the detection and analysis of posttranslational modifications of IGFBPs.

Adult↗
PubMed1998

Regulation of steroidogenesis by insulin-like growth factors (IGFs) in adult human adrenocortical cells: IGF-I and, more potently, IGF-II preferentially enhance androgen biosynthesis through interaction with the IGF-I receptor and IGF-binding proteins.

Although the effect of insulin-like growth factors (IGFs) in fetal adrenocortical cells has been investigated extensively, the role of the IGF system in the adult human adrenal gland remains unclear. In the present study we investigated the effect of recombinant human IGF-I and IGF-II on cortisol, dehydroepiandrosterone sulfate (DHEA-S) and cAMP synthesis in adult human adrenocortical cells in primary culture. Both IGFs stimulate basal as well as adrenocorticotropin (ACTH)-induced steroid secretion in a time- and dose-dependent fashion. While both IGFs (6.5 nM) induced only a moderate 2-fold increase in basal cortisol output after 48 h, the effect on basal DHEA-S secretion was significantly stronger, with a 2.7- and 3.7-fold stimulation by IGF-I and IGF-II respectively. Similarly, IGF-II enhanced ACTH-induced cortisol and DHEA-S secretion more potently than IGF-I. In dose-response experiments, the maximum stimulation of ACTH-induced DHEA-S secretion was induced by 1.6 nM IGF-I (2-fold increase) or IGF-II (2.9-fold increase), while the maximum response of cortisol secretion was elicited only at 13 nM IGF-I (2-fold increase) or IGF-II (2.5-fold increase). This resulted in a significant shift of the DHEA-S dose-response curves to the left, indicating a relative selective stimulation of androgen biosynthesis by physiologically low concentrations (0.4-3.2 nM) of IGF-II, and less potently by IGF-I. At all doses tested, the steroidogenic effect of IGF-II was significantly stronger than the effect of IGF-I. Although both IGF receptors are present in adult human adrenocortical cells, the steroidogenic effect of IGF-II is mediated through the IGF-I receptor, since [Arg54,55]IGF-II, which only binds to the IGF-I receptor, was equipotent with native IGF-II, whereas [Leu27]IGF-II, which preferentially binds to the type II IGF receptor, did not show any effect. In addition, [des1-3]IGF-I, which exhibits only minimal binding to IGFBPs, was significantly more potent than native IGF-I in stimulating adrenal steroid biosynthesis, and elicited almost the same maximum stimulatory effect as IGF-II and [des1-6]IGF-II. By Western ligand blotting of conditioned medium it was shown that adult human adrenocortical cells secrete various IGF-binding proteins (IGFBPs), which are induced differentially by treatment with ACTH. In conclusion, these results demonstrate that: (1)IGF-II stimulates basal as well as ACTH-induced DHEA-S and cortisol secretion from adult human adrenocortical cells more potently than IGF-I; (2) both IGFs predominantly stimulate androgen biosynthesis; (3) the steroidogenic effect of IGF-I and IGF-II is mediated through interaction with the IGF-I receptor; (4) the different steroidogenic potency of IGF-I and IGF-II might be explained by interaction of these ligands with locally produced IGFBPs. These data indicate that the IGF system plays an important role in the regulation of the differentiated function of adult human adrenocortical cells.

Adrenal Cortex↗
PubMed1997

[Intraoperative diagnosis of a multilocal pheochromocytoma].

Pheochromocytomas are functionally active, catecholamine-secreting tumours of chromaffin tissue. The mainstay of pharmacological therapy is preoperative treatment with oral phenoxybenzamine. This drug irreversibly alkylates alpha-1-adrenergic receptors on vascular smooth muscle and renders them nonfunctional, thereby causing vasodilatation. The duration of action of a single dose is approximately 24 h. Therefore, postoperative hypotension is a hazard of therapy with phenoxybenzamine if adequate plasma volume repletion is not provided. Prazosin, a short-acting, competitive alpha-1 blocker, has been used preoperatively, but has been criticized for its failure to adequately prevent perioperative hypertensive episodes. We report the case of a 73-year-old woman who was admitted for elective pheochromocytoma resection. Preoperative therapy with phenoxybenzamine was impossible because of the patient's refusal to take the drug. Preoperative antihypertensive preparation was therefore performed with prazosin 30 mg/24 h and metoprolol 100 mg/24 h. During the surgical preparation of the tumor, sodium nitroprusside was started at an average infusion rate of 4.1 micrograms/kg/min. After resection of the primary tumor, when the sodium nitroprusside infusion was stopped the patient exhibited an increase in systolic blood pressure (BP) up to 210 mg Hg. This hypertensive crisis was managed with sodium nitroprusside, nitroglycerin, and esmolol. A multilocular pheochromocytoma was diagnosed. Further stimuli due to tumour palpation resulted in repeated increases in BP. In this manner, two additional areas of tumour could be diagnosed by BP peaks after reduction of the sodium nitroprusside infusion. After complete resection of a total of three tumours, no further hypertensive crises occurred. The patient's postoperative course was uneventful. We conclude that in this patient presenting with an unsuspected multilocular pheochromocytoma, the lack of permanent alpha-blockade was probably helpful in allowing complete resection of all the tumours.

Adrenal Gland Neoplasms↗
PubMed1997

Interleukin-3 and interleukin-6 stimulate cortisol secretion from adult human adrenocortical cells.

Accumulating data indicate that interleukins can activate the hypothalamic-pituitary-adrenal (HPA) axis. We evaluated the effect of human recombinant interleukin-3 (IL-3) and interleukin-6 (IL-6) on cortisol secretion from adult human adrenocortical cells in primary culture. IL-3 and IL-6 (100 microg/L) equipotently stimulated basal cortisol secretion approximately 5-fold. The stimulatory effect was significant after 12 h (p<0.01) and maximum cortisol levels were induced after 48 h. In contrast to ACTH, which significantly induced cAMP levels in parallel to its steroidogenic effect, IL-3 or IL-6 had no significant effect on cAMP. Furthermore, we showed that specific inhibition of the cyclooxygenase pathway by indomethacin completely blocked the steroidogenic effect of IL-6 while the effect of IL-3 was not affected. In contrast, coincubation with nordihydroguaiaretic acid--a specific inhibitor of the lipoxygenase system--abolished IL-3 stimulated steroidogenesis but had no effect on IL-6 stimulated cortisol secretion. These findings indicate that IL-3 and IL-6 directly stimulate the steroidogenesis at the adrenal level through activation of different, cAMP-independent pathways. While the stimulatory effect of IL-6 on cortisol secretion from adult human adrenocortical cells seems to be mediated through the cyclooxygenase pathway, the effect of IL-3 on adrenocortical cortisol secretion is dependent on the lipoxygenase pathway.

Adrenal Cortex↗
PubMed1997

Sporadic pheochromocytomas are rarely associated with germline mutations in the vhl tumor suppressor gene or the ret protooncogene.

Pheochromocytoma is a tumor that may occur sporadically or may be a manifestation of a hereditary disease, such as von Hippel-Lindau disease (VHL) and multiple endocrine neoplasia (MEN) type 2. As patients with VHL or MEN type 2 are at risk to develop multiple tumors, they must be distinguished from sporadic cases. We determined the incidence of VHL and MEN type 2 among 62 German patients diagnosed with pheochromocytoma without a history of a hereditary disease. Germline analyses of the vhl gene and exons 10, 11, and 13 of the ret protooncogene were performed by PCR, single strand conformation polymorphism, enzyme digestion, or sequencing. Two patients (3%) showed vhl mutations (95% confidence interval, 1-11%). One patient showed loss of the MspI restriction site at nucleotides 712/713. Another patient had a C/T change at an intronic site that was also detected in 2 of his offspring. No mutations were detected in the ret protooncogene (97.5% confidence interval, 0-6%). In Germany, most sporadic pheochromocytomas are not due to VHL or MEN type 2. Therefore, clinical work-up in patients with pheochromocytoma without signs of hereditary disease is not recommended. However, because the costs of genetic screening are relatively low, and each index case allows optimal care for family members, molecular testing might be cost-effective.

Adolescent↗
PubMed1997

Insulin-like growth factor receptors in normal and tumorous adult human adrenocortical glands.

We have identified and characterized insulin-like growth factor (IGF)-I and IGF-II/mannose-6-phosphate (IGF-II/M6P) receptors in normal adult human adrenocortical tissue. Furthermore, we investigated the IGF-I receptor concentration and binding characteristics in benign and carcinomatous adrenocortical tumors. Membrane preparations of 14 normal adrenocortical glands showed a mean specific 125I-IGF-I binding (SB) of 5.0 +/- 0.5% and a competition by unlabeled ligands which is characteristic of the IGF-I receptor. The Scatchard analysis revealed a single class of high affinity binding sites with a dissociation constant (Kd) of 0.16 +/- 0.03 nmol/l, and a receptor concentration (RC) of 19.2 +/- 2.5 nmol/kg protein. Affinity cross-linking experiments with normal and tumorous adrenocortical tissue displayed a band at an apparent molecular mass of 135 kDa, corresponding to the size of the normal alpha-subunit of the IGF-I receptor. In agreement, 125I-IGF-II binding to normal adult human adrenocortical membranes was characteristic for the IGF-II/M6P receptor, and the Scatchard analysis revealed the presence of a single class of high affinity binding sites (SB 7.5 +/- 0.5%, RC 1137 +/- 265 nmol/kg protein, Kd 2.20 +/- 0.46 nmol/l, n = 6). The identity of the IGF-II/M6P receptor in adrenocortical tissue was further confirmed by Western blotting showing a specific band at 220 kDa. When 125I-IGF-I binding in adrenocortical hyperplasias (SB 4.1 +/- 0.4%, RC 19.6 +/- 2.0 nmol/kg protein, Kd 0.19 +/- 0.04 nmol/l, n = 4) and adenomas (SB 4.0 +/- 1.1%, RC 17.5 +/- 3.1 nmol/kg protein, Kd 0.21 +/- 0.04 nmol/l, n = 4) was compared with the 125I-IGF-I binding in normal adrenocortical tissue, similar IGF-I receptor concentration and binding kinetics were found. In contrast, three out of four hormonally active adrenocortical carcinomas showed a strongly elevated specific 125I-IGF-I binding with a 3- to 4-fold increase in IGF-I receptor concentration, as compared with normal adrenocortical tissue. This resulted in a significantly higher mean specific binding and receptor concentration in adrenocortical carcinomas, while the binding kinetics and the size of the alpha-subunit of the IGF-I receptor remained unaltered (n = 4, SB 13.8 +/- 4.2%, RC 72.2 +/- 21.3 nmol/kg protein, Kd 0.17 +/- 0.02 nmol/l). In summary, we show that intact IGF-I and IGF-II receptors are present in normal adult human adrenocortical tissue. While the abundance of the IGF-I receptor in adrenocortical hyperplasias and adenomas was similar to normal tissue, a strong overexpression of the intact IGF-I receptor was found in three out of four adrenocortical carcinomas.

Adenoma↗
PubMed1996

Mutations in the VHL tumor suppressor gene and associated lesions in families with von Hippel-Lindau disease from central Europe.

von Hippel-Lindau (VHL) disease is a dominantly inherited familial cancer syndrome predisposing to retinal, cerebellar and spinal hemangioblastoma, renal cell carcinoma (RCC), pheochromocytoma and pancreatic tumors. Clinically two types of the disease can be distinguished: VHL type 1 (without pheochromocytoma) and VHL type 2 (with pheochromocytoma). We report VHL germline mutations and trends in phenotypic variation in families from central Europe. We identified 28 mutations in 53/65 (81.5%) families with 18 (64%) mutations being unique to this population. Whereas types and distribution of mutations as well as a strong correlation of missense mutations with the VHL 2 phenotype were similar to those identified in other populations, these families have provided new insights into the molecular basis for variability in the VHL 2 phenotype. Seven different missense mutations in exons 1 and 3 varied in their biological consequences from a minimal VHL 2 phenotype with pheochromocytoma only to a full VHL 2 phenotype with RCC and pancreatic lesion. These findings contribute to a better understanding of the fundamental mechanisms of VHL disease and its phenotypic variability. Further, we have provided rapid VHL screening for the families in central Europe, which has resulted in improved diagnosis and clinical management.

Base Sequence↗
PubMed1996

Isolated familial pheochromocytoma as a variant of von Hippel-Lindau disease.

Inherited pheochromocytomas are often part of familial syndromes, especially multiple endocrine neoplasia type 2 (MEN 2), retinal cerebellar hemangioblastomatosis [von Hippel-Lindau (vHL) disease] or neurofibromatosis type 1. It is not clear whether isolated familial pheochromocytoma exists as a separate clinical entity. In a family with pheochromocytomas in three generations and with at least seven affected members, we investigated by clinical and genetic analyses the presence or absence of associated conditions. The clinical investigations included ophthalmological and radiological studies for von Hippel-Lindau disease (magnetic resonance imaging of the brain, computed tomography of the abdomen, and direct ophthalmoscopy after mydriasis) and annual calcitonin stimulation tests for C cell disease in five members who agreed to regular follow-up. Besides the pheochromocytomas (so far, these have been multiple in five of seven individuals) no definite second associated condition was found. Genetic analysis did not identify any MEN 2-specific RET protooncogene point mutations (which are present in 97% of MEN 2a families). However, despite the complete absence of other clinical manifestations of the vHL disease (besides pheochromocytomas), a previously undescribed germline missense mutation in the vHL tumor suppressor gene was found (C775G transversion with a predicted substitution of a leucine by a valine at codon 259 in the putative vHL protein). We conclude that in this family the sole occurrence of pheochromocytoma is a variant of vHL disease.

Adrenal Gland Neoplasms↗
PubMed1995

Insulin-like growth factor II (IGF-II) is more potent than IGF-I in stimulating cortisol secretion from cultured bovine adrenocortical cells: interaction with the IGF-I receptor and IGF-binding proteins.

Although the stimulating effect of insulin-like growth factor I (IGF-I) on adrenal steroidogenesis has been well established, the role of IGF-II in the adult adrenal gland remains unknown. We, therefore, investigated the effect of recombinant human IGF-II on cortisol and cAMP synthesis from adult bovine adrenocortical cells. IGF-II, time and dose dependently, stimulated basal cortisol secretion maximally 3-fold. In combination with ACTH, IGF-II (13 nM) synergistically increased cortisol secretion from 1-fold (10(-8) M ACTH) to 28-fold of untreated control levels. In contrast, IGF-I at equimolar concentrations did not show an effect on basal cortisol secretion, and in combination with ACTH elicited a significant weaker stimulatory effect than IGF-II (22-fold increase). The synergistic effect of IGF-II on ACTH-promoted cortisol secretion was paralleled by accumulation of cAMP in the culture medium. Although both IGF receptors are present in adult bovine adrenocortical cells, the effect of IGF-II seems to be mediated through interaction with the IGF-I receptor, as [Arg54,55]IGF-II, which only binds to the IGF-I receptor, was equipotent to native IGF-II, whereas [Leu27]IGF-II, which preferentially binds to the type II IGF receptor, did not show any effect. By Western ligand blotting, four different molecular forms of IGF-binding proteins (IGFBPs) were identified in conditioned medium of bovine adrenocortical cells with apparent molecular masses of 39-44, 34, 29, and 24 kilodaltons. ACTH treatment increased the abundance of all binding proteins, on the average, 2.3-fold, except for the 29-kDa band, which was predominantly induced 6.8-fold. Additionally, [des1-3]IGF-I, a truncated IGF variant that exhibits only minimal binding to IGFBPs, was significant more potent than IGF-I and elicited the same maximum stimulatory effect on cortisol secretion as IGF-II and [des1-6]IGF-II. In conclusion, these results demonstrate that 1) IGF-II stimulates basal as well as ACTH-induced cortisol secretion from bovine adrenocortical cells more potently than IGF-I; 2) this effect is mediated through interaction of IGF-II with the IGF-I receptor; 3) bovine adrenocortical cells synthesize various IGFBPs that are induced differentially by ACTH; and 4) IGFBPs apparently play a modulatory role in IGF-induced stimulation of adrenal steroidogenesis. Therefore, bovine adult adrenocortical cells provide a useful tissue culture model in which the interactions among locally produced IGFs, IGFBPs, and the IGF-I receptor can be evaluated.

Adrenal Cortex↗