New England Deaconess Hospital Research Articles

Leukocyte common-antigen-related tyrosine phosphatase receptor: altered expression of mRNA and protein in the New England Deaconess Hospital rat line exhibiting spontaneous pheochromocytoma

Regulation of cell proliferation by protein tyrosine phosphatases (PTPs) suggests that PTPs are important tumor suppressor genes. The gene encoding the leukocyte common-antigen-related (LAR) PTP receptor maps to chromosome 1p32-33, a region in which loss of heterozygosity is associated with human pheochromocytoma and other neuroectodermal tumors. The rat pheochromocytoma PC12 cell line was originally derived from the transplantable P259 tumor originating from the New England Deaconess Hospital (NEDH) line of Wistar inbred rats. Compared with their Wistar counterparts, 1-2-year-old NEDH rats exhibit a high incidence of spontaneous pheochromocytomas. This study investigates whether levels of LAR transcripts and protein are altered in NEDH adrenal tissue prior to tumor onset. In addition, alternative splicing of an LAR extracellular domain [LAR alternatively spliced element-c (LASE-c)], regulating LAR interaction with extracellular matrix components, was examined. These changes in LAR expression and alternative splicing were hypothesized to be more pronounced in tumor tissue and PC12 cells. Northern blot analysis demonstrated the presence of the approximately 5 kb LAR transcript in all cell lines examined, except PC12. In adrenal medulla tissue harvested from 2-3-month-old rats, LAR approximately 8 and approximately 5 kb transcript expression was decreased in NEDH compared with Wistar samples. RT-PCR demonstrated increased splicing of the LASE-c 27 bp alternatively spliced insert in the LAR extracellular domain in NEDH adrenal medulla tissue. Even greater LASE-c splicing was detected in adrenal medulla tumor tissue derived from 12-month-old NEDH rats and in PC12 cells. Western blot analysis demonstrated decreased levels of LAR protein and increased levels of LASE-c containing LAR protein isoforms in NEDH adrenal medulla tissue. These studies demonstrate that patterns of altered LAR expression present in PC12 cells and in pheochromocytoma tumor tissue are also present in adrenal tissue predisposed to a high incidence of spontaneous pheochromocytoma.

Association between sucrase-isomaltase and p53 expression in colorectal cancer.

Sucrase-isomaltase (SI) is a tissue-based phenotypic marker that is an independent prognostic factor in colorectal cancer (CRC). DF3 and galectin 3 are two other tissue-based markers that are upregulated during neoplastic transformation. Because p53 mutations are acquired during neoplastic progression, we reasoned that alterations in SI and p53 may be associated despite an apparent lack of biological interaction. Paraffin sections from 183 patients who underwent surgery at New England Deaconess Hospital (NEDH) between 1965 and 1977 were analyzed first by immunohistochemistry (IHC) for the expression of the markers SI, DF3, and galectin 3, which were scored as absent or present. Paraffin sections from a second group of 59 patients who underwent surgery at NEDH between 1985 and 1992 were analyzed by IHC for the expression of p53 as well as SI, DF3, and galectin 3. p53 nuclear staining was scored as absent or present. Previous work has shown that p53 is mutated in all cells with nuclear staining and in 10% of tumors that are unstained. SI expression was not associated with the expression of either DF3 or galectin 3, and neither DF3 nor galectin 3 were prognostic factors in CRC. None of the phenotypic markers were associated with any of the clinicopathologic variables. However, 21 of 24 p53-positive cases (88%) expressed SI, whereas 15 of 35 p53-negative cases (43%) were also SI negative (p = 0.02, Fisher exact test). p53 expression was not associated with expression of DF3 or galectin 3. SI expression and p53 mutation are associated significantly in CRC. Although the mechanism underlying such an association in presently unknown, the association may define a subset of patients with a worse prognosis.

S-4-5 - Pheochromocytoma (PHEO)-harboring New England Deaconess Hospital (NEDH) rats: A model for hypertension and in vivo adrenoceptor desensitization.

S-4-5 - Pheochromocytoma (PHEO)-harboring New England Deaconess Hospital (NEDH) rats: A model for hypertension and in vivo adrenoceptor desensitization.

Hyperinsulinaemia causes a preferential increase in hepatic p4501A2 activity

Male NEDH (New England Deaconess Hospital) rats were transplanted with a radiationinduced tumour from a donor male rat and were killed 18 days following transplantation. At the time of killing the insulinoma-bearing animals were severely hypoglycaemic but plasma ketone levels were normal. Insulinoma-bearing animals exhibited higher hepatic O-deethylation of ethoxyresorufin and N-demethylation of ethylmorphine activities when compared to control animals. Similarly, hepatic microsomal preparations from insulinoma-bearing rats were more efficient than control animals in converting the promutagen 2-amino-6-methyldipyrido[1,2- a:3′,2′']imidazole (Glu-P-1) to mutagenic intermediates in the Ames test. Immunoblot analysis employing polyclonal antibodies against the P4501A and P453A families revealed that insulinoma-bearing rats had higher hepatic P4501A2 apoprotein levels. No major differences in P4503A1 apoprotein levels between insulinoma-bearing and control rats were noted. Subcutaneous administration of insulin to male Wistar rats gave rise to a modest increase in ethoxyresorufin O-deethylase activity and in the ability to activate Glu-P-1 to mutagens in the Ames test. Immunoblot analysis revealed an increase in hepatic P4501A2 apoprotein levels following the treatment with insulin. It is concluded that insulinoma-bearing rats display high P4501A2 activity and the hyperinsulinaemia that characterize this condition is responsible for the effect. Moreover, administration of insulin to other strains of rat, such as Wistar, also enhances P4501A2 activity, presumably as a result of hyperinsulinaemia.

Desensitization of α-Adrenergic Receptor-Mediated Smooth Muscle Contraction

Desensitization of alpha-adrenergic receptor-mediated smooth muscle contraction occur in aortas from New England Deaconess Hospital (NEDH) rats harboring pheochromocytoma (PHEO) and following chronic exposure to the alpha-adrenergic agonist phenylephrine in vitro. Endothelium is known to release an endothelial cell-derived relaxing factor that promotes smooth muscle relaxation. We wondered if the endothelium might contribute to the desensitization of contraction. The role of the endothelium in desensitization was studied using aortic rings with endothelium [E(+)] and with endothelium removed [E(-)]. Maximal phenylephrine (PE)-induced contraction (Emax) for E(+) was 1.7 +/- 0.3 g in controls and 0.4 +/- 0.1 g in PHEO (p less than 0.001), demonstrating desensitization; however, for E(-), Emax was 2.4 +/- 0.2 g in PHEO vs. 2.5 +/- 0.2 g in controls, demonstrating restoration of maximal contraction when the endothelium was removed. However, sensitivity [-log EC50(M)] to PE in E(-) remained significantly lower in PHEO compared to controls (6.94 +/- 0.12 vs. 7.51 +/- 0.14, respectively, p less than 0.001). Similarly, in aortic ring segments desensitized in vitro with phenylephrine, the maximal contraction in phenylephrine-exposed aortas was 60% of that seen in controls. Removal of the endothelium from the vessels pretreated with phenylephrine fully restored the maximal response and sensitivity of these vessels. Treatment of desensitized vessels with hemoglobin (5 x 10(-5) M) restored the maximal contraction and sensitivity to phenylephrine. When the endothelium was removed prior to chronic exposure to phenylephrine, the sensitivity to phenylephrine decreased while the Emax remained similar to controls.(ABSTRACT TRUNCATED AT 250 WORDS)

Potentially curative surgery of colon cancer: patterns of failure and survival.

In an effort to determine the patterns of failure and survival of colon cancer, a retrospective review of 294 patients who underwent potentially curative surgery at the New England Deaconess Hospital (NEDH) was performed. For the entire group, the 5-year crude survival rate was 68% and the actuarial rate was 80%. Survival decreased with increasing bowel wall penetration by tumor and the presence of lymph node metastasis. Although survival varied with the tumor site, none of the differences was statistically significant. Other variables, including the grade of adenocarcinoma, size, and the type of surgery had a significant impact on survival. Patterns of failure, expressed as the actuarial incidence of first diagnosed failure at 5 years, were examined by stage and site. There was a trend toward increased failure with increasing bowel wall penetration by tumor and the presence of lymph node metastasis. Abdominal failure, either as the only site or as a component of failure, was the most common type of failure. When compared by site, patients with cecal carcinoma had a significantly lower incidence of local and distant failure than patients with disease in other selected sites. No differences in patterns of failure were seen in patients with carcinomas in the mobile sections of the colon compared with those who had disease arising in the nonmobile sections of the colon. These data may be useful in identifying those patients who might benefit most from adjuvant therapy.

Potentially curative surgery of colon cancer: the influence of blood vessel invasion.

A number of series have examined the influence of blood vessel invasion (BVI) by tumor on survival of patients with colorectal cancer; however, there are little data available regarding its influence on patterns of failure. In an effort to determine the influence of BVI on the patterns of failure and survival in colon cancer, a retrospective review of 294 patients who underwent potentially curative surgery at the New England Deaconess Hospital (NEDH) was performed. Patients whose tumors had BVI experienced a significant decrease in the 5-year actuarial survival rate. BVI had little impact on the patterns of failure in stage B2 disease, but a significant increase in total failure and local failure (as a component of failure) occurred in stage C2. However, when examined by proportional hazards analysis, BVI was found not to be an independent prognostic variable. For patients with stage C2 tumors, which are also BVI+, radiation therapy to the tumor bed might play a contributory role in overall management.

Defective regulation of glucokinase in rat pancreatic islet cell tumours.

The role of glucokinase in the regulation of insulin secretion was examined in normal rat pancreatic islets and in chemically- and radiation-induced rat pancreatic B-cell tumours which show an impaired insulin secretory response to glucose. In normal rats glucokinase activity in cytoplasmic fractions of pancreatic islets was decreased with the duration of fasting and increased by refeeding or insulin administration. This observation is consistent with the induction of glucokinase by insulin. Hexokinase activity was only slightly reduced during fasting. Glucokinase activity decreased in cytoplasmic fractions of streptozotocin-nicotinamide-induced rat pancreatic islet cell tumours. Glucokinase activity contributed about 75% to the total glucose phosphorylation capacity in cytoplasmic fractions of normal pancreatic islets and of small (less than 1 mg) streptozotocin-nicotinamide-tumours. This proportion decreased to about 20% in the large streptozotocin-nicotinamide tumours. Glucokinase activity in cytoplasmic fractions of transplantable radiation-induced NEDH (New England Deaconess Hospital) rat B-cell tumours was seven times lower than in normal pancreatic islets and contributed only 15% to the total glucose phosphorylation capacity. In contrast, hexokinase activity of the NEDH tumour B-cells was 2.5 times higher than normal. Decreased glucokinase activity in the chemically- and radiation-induced tumour B-cells appears to result from a loss of the ability of insulin to induce this enzyme and may explain the lack of insulin secretory responsiveness of these tumour B-cells.

Effect of pheochromocytoma and hypophysectomy on blood pressure and catecholamines in NEDH rats.

The New England Deaconess Hospital (NEDH) rat provides a valuable model with which to study pheochromocytoma (P); 59% of male rats 700 to 900 days old and 81% of those 900 days or older developed spontaneous P. One transplantable P (P259), when implanted into other NEDH rats, markedly increased plasma norepinephrine and dopamine as well as blood pressure, and usually caused death within 4 weeks. Even without P, about 83% of NEDH rats became hypertensive by 131/2 weeks of age and remained moderately hypertensive until 2 years of age when some animals developed spontaneous P and hypertension became severe. Whether a common mechanism is responsible for early appearance of hypertension and later development of P remains to be determined. Hypophysectomized NEDH rats remained normotensive or slightly hypotensive despite marked elevations of plasma norepinephrine and dopamine caused by P259 implantation; furthermore, survival was prolonged to 3 months. Catecholamine concentrations in plasma and RBC were usually quite similar, indicating that red blood cells play a significant role in inactivating circulating catecholamines. Unlike the normal adrenal, P259 in NEDH rats contains mainly norepinephrine and dopamine with little epinephrine; it appears that P259 is deficient in the enzyme phenylethanolamine-N-methyltransferase (PNMT), which converts norepinephrine to epinephrine. Why hypophysectomy prevents hypertension and prolongs life in rats with P259 implants is unclear; adrenal cortical and thyroid deficiency may play a role. Preliminary observations indicate that hypophysectomy can prevent spontaneous development of P in NEDH rats.

Studies of a Transplantable Rat Pheochromocytoma

A transplantable rat pheochromocytoma in New England Deaconess Hospital (NEDH) was first described by WARREN in 1972. It is characterized by the documented association with systolic hypertension and the known presence of increased urinary metanephrines and vanilmadelic acid in tumor-bearing animals. The present report describes features of the transplantable tumor, our laboratories have noted, in five tumor transplantations starting in 1974. Tumor-bearing animals survive 49 +/- 5 days and die much sooner than aging, non-tumor-bearing litter mates. Gross measurements confirm the rapid growth of the primary tumor, although at autopsy, histologically proven metastatic foci are rarely seen. Polycythemia with or without increased erythropoietin (ESF) levels were not detected. Electronmicroscopic studies confirmed the presence in tumor tissue of the previously described intracytoplasmic granules. Further studies on this endocrine-associated transplantable tumor are warranted and feasible.

RESULTS OF GALLBLADDER SURGERY IN DIABETES MELLITUS

St. Louis, Mo. FROM THE GEORGE F. BAKER CLINIC, NEW ENGLAND DEACONESS HOSPITAL, BOSTON, MASS., ELLIOTT P. JOSLIN, M.D., MEDICAL DIRECTOR * Fellow in Medicine, George F. Baker Clinic, New England Deaconess Hospital, Boston, Mass., July, 1941-July, 1942.

RESULTS OF GALLBLADDER SURGERY IN DIABETES MELLITUS

St. Louis, Mo. FROM THE GEORGE F. BAKER CLINIC, NEW ENGLAND DEACONESS HOSPITAL, BOSTON, MASS., ELLIOTT P. JOSLIN, M.D., MEDICAL DIRECTOR * Fellow in Medicine, George F. Baker Clinic, New England Deaconess Hospital, Boston, Mass., July, 1941-July, 1942.

DIABETIC GANGRENE

In this paper I shall try to give a general picture of the diabetic patient with gangrene, to describe the organization for and the principles of management of cases of diabetic gangrene at the New England Deaconess Hospital and possibly, through a study of my results, to indicate how I have arrived at my present conception of the operative treatment of this condition. I do not wish to add to the many reports on the details of management of diabetic gangrene or to enter into any controversy as to the indications for amputation or the level at which it should be done. ORGANIZATION FOR THE CARE OF SURGICAL DIABETIC PATIENTS AT THE NEW ENGLAND DEACONESS HOSPITAL I did my first amputation for gangrene—a closed amputation through the upper third of the lower part of the leg—in May 1923. Insulin had been given to the first patient in the New England