Enzyme-altered foci in colons of carcinogen-treated rats.

The distal colon and rectum from male F344 rats treated with 15 mg/kg 1,2-dimethylhydrazine.2HCl (DMH) for 20 weeks were analyzed for focal areas of enzyme alteration. Tissues were embedded in methacrylate at 4 degrees C and cut in 2- to 4-micron serial sections. In DMH-treated rats, 8.8 +/- 2.4 foci/cm2 of examined mucosa were observed at 20 weeks and 7.7 +/- 1.1 foci/cm2 at 31 to 52 weeks, compared with 1.2 +/- 0.6 foci/cm2 in control rats (P = 0.01). The number of foci at 31 to 52 weeks compared with 20 weeks did not change significantly, but the area of altered rectal mucosa increased from 0.22 +/- 0.2% at 20 weeks to 1.47 +/- 0.6% at 31 to 52 weeks (P = 0.051). Most foci had decreased N-acetyl-beta-D-glucosaminidase, alpha-naphthyl butyrate esterase, and mucin in epithelial cells and increased gamma-glutamyl transpeptidase in the stroma. Morphologically, the foci varied from normal to overtly dysplastic. Grossly, tumors were identified in 5 of 20 DMH-treated rats killed at 31 to 52 weeks but not in 12 DMH-treated rats killed at 20 weeks or 30 control rats killed at 20 to 52 weeks. These data suggest but do not establish that enzyme-altered foci are putative preneoplastic lesions in the colon.

Aberrant crypts: putative preneoplastic foci in human colonic mucosa.

Aberrant crypts were identified for the first time in whole-mount preparations of normal-appearing human colonic mucosa after staining with methylene blue. The foci of aberrant crypts varied from single altered glands to plaques of greater than 30 crypts. The mean proportion of colonic mucosa altered and the number of foci with aberrant crypts per cm2 of colonic mucosa were (a) higher in patients with colon cancer than in patients without colon cancer or predisposing conditions and (b) highest in our single case of Gardner's syndrome. Aberrant crypts are postulated to be the earliest identifiable potential precursors of colon cancer. Analysis of aberrant crypts may facilitate the study of the early pathological and molecular changes that precede colon cancer.

Normalization of insulin responses to glucose by overnight infusion of glucagon-like peptide 1 (7-36) amide in patients with NIDDM.

Glucagon-like peptide 1 (GLP-1) is a natural enteric incretin hormone, which is a potent insulin secretogogue in vitro and in vivo in humans. Its effects on overnight glucose concentrations and the specific phases of insulin response to glucose and nonglucose secretogogues in subjects with NIDDM are not known. We compared the effects of overnight intravenous infusion of GLP-1 (7-36) amide with saline infusion, on overnight plasma concentrations of glucose, insulin, and glucagon in eight subjects with NIDDM. The effects on basal (fasting) beta-cell function and insulin sensitivity were assessed using homeostasis model assessment (HOMA) and compared with seven age- and weight-matched nondiabetic control subjects. The GLP-1 infusion was continued, and the first- and second-phase insulin responses to a 2-h 13 mmol/l hyperglycemic clamp and the insulin response to a subsequent bolus of the nonglucose secretogogue, arginine, were measured. These were compared with similar measurements recorded after the overnight saline infusion and in the control subjects who were not receiving GLP-1. The effects on stimulated beta-cell function of lowering plasma glucose per se were assessed by a separate overnight infusion of soluble insulin, the rate of which was adjusted to mimic the blood glucose profile achieved with GLP-1. Infusion of GLP-1 resulted in significant lowering of overnight plasma glucose concentrations compared with saline, with mean postabsorptive glucose concentrations (2400-0800) of 5.6 +/- 0.8 and 7.8 +/- 1.4 mmol/l, respectively (P < 0.0002). Basal beta-cell function assessed by HOMA was improved from geometric mean (1 SD range), 45% beta (24-85) to 91% beta (55-151) by GLP-1 (P < 0.0004). First-phase incremental insulin response to glucose was improved by GLP-1 from 8 pmol/l (-8-33) to 116 pmol/l (12-438) (P < 0.005), second-phase insulin response to glucose from 136 pmol/l (53-352) to 1,156 pmol/l (357-3,748) (P < 0.0002), and incremental insulin response to arginine from 443 pmol/l (172-1,144) to 811 pmol/l (272-2,417) (P < 0.002). All responses on GLP-1 were not significantly different from nondiabetic control subjects. Reduction of overnight glucose by exogenous insulin did not improve any of the phases of stimulated beta-cell function. Prolonged intravenous infusion of GLP-1 thus significantly lowered overnight glucose concentrations in subjects with NIDDM and improved both basal and stimulated beta-cell function to nondiabetic levels. It may prove to be a useful agent in the reduction of hyperglycemia in NIDDM.

Relative hyperproinsulinemia of NIDDM persists despite the reduction of hyperglycemia with insulin or sulfonylurea therapy.

Subjects with NIDDM have increased plasma proinsulin concentrations, compared with nondiabetic subjects, both in absolute terms and as a proportion of circulating insulin-like molecules. It remains uncertain whether this reflects a primary beta-cell defect in proinsulin processing or is secondary to hyperglycemia. We addressed this question by assessing the effects of reducing hyperglycemia on relative hyperproinsulinemia in subjects with NIDDM. Eight subjects with NIDDM underwent three 8-week periods in a randomized crossover design of therapy with diet alone, sulfonylurea (gliclazide), or insulin (ultralente). The effects on beta-cell peptide concentrations were assessed 1) fasting, 2) in response to hyperglycemic clamping, and 3) in response to an injection of the nonglucose secretogogue arginine and compared with measurements in seven nondiabetic control subjects. Both sulfonylurea and insulin therapy substantially reduced fasting plasma glucose and glycosylated hemoglobin (HbA1e) concentrations, compared with diet therapy alone. The diabetic subjects on diet therapy had relative hyperproinsulinemia, assessed relative to C-peptide concentrations, fasting and in response to hyperglycemic clamping and arginine, compared with control subjects. Neither sulfonylurea nor insulin therapy altered the relative hyperproinsulinemia. Insulin therapy reduced fasting proinsulin concentrations from geometric mean 29.4 (1 SD range, 14.6-59.0) pmol/l on diet therapy to 18.7 (7.3-48.1) pmol/l (P < 0.05). A similar trend was evident with fasting C-peptide concentrations with a reduction from 0.9 (0.6-1.4) nmol/l on diet therapy to 0.6 (0.4-0.9) nmol/l (P = 0.06), so that the relative hyperproinsulinemia, assessed as the ratio of fasting proinsulin to C-peptide, was unchanged by insulin. Similarly, insulin therapy failed to reduce the ratio of proinsulin to C-peptide concentrations in response to a hyperglycemic clamp and in the acute incremental response to arginine. Failure to improve the relative hyperproinsulinemia of NIDDM, despite significant reduction of hyperglycemia with exogenous insulin therapy, supports the hypothesis that relative hyperproinsulinemia in NIDDM is a reflection of a primary beta-cell defect rather than being secondary to hyperglycemia.

Availability of type II diabetic families for detection of diabetes susceptibility genes.

Type II diabetes is a familial disorder, as evidenced by the increased prevalence in monozygotic cotwins and first-degree relatives of affected subjects; however, its genetic etiology is largely unknown. Well-characterized pedigrees are an essential resource for the study of susceptibility genes for type II diabetes. This study describes a 5-yr search for type II diabetic families in Oxfordshire, U.K. We interviewed 950 type II diabetic subjects concerning the availability of first-degree relatives; 127 Caucasian families ascertained through a proband with type II diabetes were studied, and 589 first-degree relatives were characterized. Three large pedigrees with maturity-onset diabetes of the young, and 8 multiplex multigenerational type II diabetic pedigrees were identified. We identified 12 sib-pairs in which both siblings had type II diabetes; however, only 7 sib-pairs had both parents alive, and 2 of these had both parents affected. If one also considers one sib having diabetes and one sib having glucose intolerance as being an affected sib-pair, we identified 30 sib-pairs of which 7 had both parents affected and probably had bilineal inheritance. We identified 76 complete nuclear families with both parents and offspring available for study, but only 6 were of optimal structure for linkage analysis. In conclusion, multiplex pedigrees and type II diabetic sib-pairs with living parents are uncommon, and their ascertainment requires a substantial investment of resources. Large-scale collaborative multicenter initiatives would be needed to collect a large resource of family material for the study of susceptibility genes for type II diabetes.

Linkage analysis of glucokinase gene with NIDDM in Caucasian pedigrees.

NIDDM has a strong genetic component, as evidenced by the high level of concordance between identical twins. The nature of the genetic predisposition has remained largely unknown. Recently, the glucokinase gene locus on chromosome 7p has been shown to be linked to a subtype of NIDDM known as MODY in French and British pedigrees, and glucokinase mutations have been identified. To study the relationship between the glucokinase gene and NIDDM, we performed a linkage analysis in 12 Caucasian pedigrees ascertained through a proband with classical NIDDM. The LINKAGE program was used under four models, including autosomal dominant and recessive, with individuals with glucose intolerance counted as either affected or of unknown status. Linkage was significantly rejected with the dominant models (LOD scores -4.65, -4.25), and was unlikely with the recessive model when glucose intolerance was considered as affected (LOD score -1.38). These findings suggest that mutations in or near the glucokinase gene are unlikely to be the major cause of the inherited predisposition to NIDDM in Caucasian pedigrees, but do not exclude a role for this locus with a polygenic model, or a major role in some pedigrees.

Changes in amylin and amylin-like peptide concentrations and beta-cell function in response to sulfonylurea or insulin therapy in NIDDM.

OBJECTIVE: Amylin, a secretory peptide of beta-cells, is the constituent peptide of islet amyloid, which is characteristic of NIDDM, and changes in amylin secretion in response to therapies may influence the rate of production of islet amyloid. The primary objective of this study was to determine whether therapy with sulfonylurea or basal insulin in NIDDM would alter amylin secretion in a way that might affect the formation of islet amyloid. RESEARCH DESIGN AND METHODS: In a randomized crossover design, eight subjects with NIDDM underwent three 8-week periods of therapy with diet alone, sulfonylurea, or exogenous basal insulin, with evaluation of amylin, amylin-like peptide (ALP), and glucose and C-peptide concentrations, both during fasting and after a standard breakfast. Changes in beta-cell function (% beta) were assessed, in the basal state by homeostasis model assessment (HOMA) and in the stimulated state by hyperglycemic clamps. Seven nondiabetic control subjects each underwent a meal profile and hyperglycemic clamp. RESULTS: Both sulfonylurea and insulin therapy reduced basal glucose concentrations compared with diet alone, but neither reduced the increased postprandial glucose increments. Both sulfonylurea and insulin therapy increased basal % beta, assessed by HOMA, but only sulfonylurea increased the second-phase C-peptide responses to the hyperglycemic clamp. Sulfonylurea increased time-averaged mean postprandial amylin and ALP concentrations compared with diet alone (geometric mean [1-SD range] for amylin, 4.9 [2.0-11.8] vs. 3.0 [1.4-6.2] pmol/l, P = 0.003; for ALP, 16.4 [8.5-31.7] vs. 10.1 [4.9-20.8] pmol/l, P = 0.001). Insulin therapy reduced basal ALP concentrations compared with diet alone (2.9 [1.5-5.6] vs. 6.0 [2.6-13.6] pmol/l, P = 0.03), but had no effect on postprandial concentrations of amylin (3.0 [1.3-6.5] pmol/l) or ALP (10.0 [5.5-18.1] pmol/l). CONCLUSIONS: By increasing postprandial concentrations of the constituent peptides of islet amyloid, sulfonylurea therapy might increase the rate of deposition of islet amyloid and thereby accelerate the decline of % beta in NIDDM, compared with diet therapy alone.

Role of radiotherapy in sarcoma of the breast--a retrospective review of the M.D. Anderson experience.

BACKGROUND: The role of adjuvant radiotherapy for sarcoma of the breast, based on local extension of disease and patterns of failure, remains undefined because of the rarity of the disease presentation. METHODS: Fifty-nine cases of soft tissue sarcoma of the breast were retrospectively reviewed. Cystosarcoma phylloides was excluded from analysis. Surgical intervention consisted of segmental resection (n = 16) or mastectomy (n = 38); five patients underwent excisional biopsy. Adjuvant radiotherapy was administered in four patients following segmental resection and in 13 patients after mastectomy. Doses totaled 50 Gy in the majority of patients, and conventional criteria and radiotherapy techniques for adjuvant breast irradiation were used. RESULTS: None of the dissected axillary nodes contained metastatic tumor on pathologic review. Patterns of failure were evaluated. Tumor size (P < 0.03) and surgical margins (P < 0.002) were predictive of local failure (LF). Due to limited patient numbers, no statistical significance was identified with any treatment modality. Following mastectomy alone, LF occurred in 13 patients (34%) versus the 13% rate of LF with mastectomy and radiotherapy (P = NS). Distant metastases developed concurrently with the two local failures in the group that underwent mastectomy and radiation. After segmental mastectomy, LF occurred in 3 cases (25%) concurrent with distant metastases: no LF were noted after segmental mastectomy and radiation (P = 0.27). For all treatment groups, local recurrences were characterized as multiple and involved the chest wall. Local failure occurred in 60% of patients with positive surgical margins who did not receive adjuvant irradiation. Irrespective of surgical margins, over 75% of local recurrences developed among patients treated by surgery alone. CONCLUSIONS: The role of radiotherapy for breast sarcoma remains undefined due to the rarity of this disease presentation. This retrospective review failed to demonstrate a statistical benefit for the administration of adjuvant irradiation in sarcoma of the breast, probably because of limited patient numbers. Because large tumor size and positive surgical margins incur a higher risk for LF, radiotherapy is probably indicated in these cases. Axillary dissection obligates the radiotherapist to treat the axilla in order to include all tissues in the surgical bed, and should be avoided to reduce potential treatment related morbidity. Established therapeutic principles and techniques used for both soft tissue sarcoma and breast cancer should continue to be applied.

Parallel changes of proinsulin and islet amyloid polypeptide in glucose intolerance.

Elevated proinsulin secretion and islet amyloid deposition are both features of Type 2 diabetes but their relationship to beta-cell dysfunction is unknown. To determine if islet amyloid polypeptide (IAPP) secretion is disproportionate with other beta-cell products at any stage of glucose intolerance, 116 subjects were studied. Non-diabetic subjects with equivalent body mass index (BMI) were assigned to three groups, (i) normal fasting glucose, fpg<5.5 mmol l(-1); (ii) intermediate fasting glucose, fpg> or =5.5<6.15 mmol l(-1); (iii) impaired fasting glucose (IFG), fpg> or =6.1<7.0 mmol l(-1). Diabetic subjects were divided according to therapy (9 diet, 19 tablet, and 11 insulin). IAPP, C-peptide and proinsulin were measured fasting and at the end of a 1-h glucose infusion. Fasting C-peptide, IAPP and proinsulin were significantly elevated in the IFG group compared with the other non-diabetic groups (P<0.02); fasting IAPP/C-peptide and proinsulin/C-peptide were 1-2% in all non-diabetic groups. Fasting and 1-h proinsulin and proinsulin/C-peptide were higher in diabetic compared with non-diabetic subjects (P<0.01). IAPP and IAPP/C-peptide in diabetic groups were similar to that in non-diabetic subjects but reduced in the insulin-treated group (P<0.01). Proinsulin was disproportionately increased compared with C-peptide and IAPP in Type 2 diabetes particularly in severe beta-cell failure implying more than one concurrent beta-cell pathology.

Hyperglycaemic siblings of Type II (non-insulin-dependent) diabetic patients have increased PAI-1, central obesity and insulin resistance compared with their paired normoglycaemic sibling.

AIMS/HYPOTHESIS: First-degree relatives of Type II (non-insulin-dependent) diabetic patients in cross-sectional studies have increased insulin resistance, associated cardiovascular risk factors and abnormalities of fibrinolysis and coagulation. To minimise between-family genetic and environmental confounders, we investigated within-family relationships between early hyperglycaemia and risk factors. METHODS: Thirteen age and gender matched sibling pairs of Type II (non-insulin-dependent) diabetic patients, one hyperglycaemic, one normoglycaemic (fasting plasma glucose at screening 6.0-7.7 mmol.l(-1) and < 6.0 mmol.l(-1), respectively) were assessed for plasminogen activator inhibitor antigen (PAI-1), tissue plasminogen activator antigen (t-PA), fibrinogen, Factor VII and Factor VIII/von Willebrand factor antigen. Fasting lipid profiles, blood pressure and HOMA insulin sensitivity (%S) were also measured in siblings and in matched subjects without family history of diabetes. RESULTS: Hyperglycaemic and normoglycaemic siblings (7 female, 6 male) were aged, mean (SD) 56.8 (8.7) and 55.8 (8.4) years. Hyperglycaemic siblings had increased PAI-1 antigen, geometric mean (i.q.r.): 26.3 (15.1-45.6) vs 11.1 (2.1-23.3) ng/ml, p=0.0002, similar t-PA antigen, mean (SD) 9.5 (4.3) vs 7.4 (2.5) ng/ml, p=0.2 and fibrinogen 2.2 (0.3) vs 2.3 (0.6) g/l, p=0.5, and reduced %S 66.3 (30.5) vs 82.9 (25), p=0.04. PAI-1 correlated negatively with %S ( r=-0.55, p=0.005). No significant differences were found in blood pressure or fasting lipids. CONCLUSION/INTERPRETATION: A minor increase in plasma glucose in non-diabetic sibling pairs of Type II (non-insulin-dependent) diabetic patients was associated with reduced insulin sensitivity, increased central adiposity and a doubling of PAI-1 antigen concentration, suggesting impaired fibrinolysis. It is possible that this could contribute to increased cardiovascular risk in these subjects.

Near-normalisation of diurnal glucose concentrations by continuous administration of glucagon-like peptide-1 (GLP-1) in subjects with NIDDM.

The gut hormone, glucagon-like peptide-1 (GLP-1) is a potent insulin secretogogue with potential as a therapy for non-insulin-dependent diabetes mellitus (NIDDM). GLP-1 has been shown to reduce glucose concentrations, both basally, and, independently, in response to a single meal. For it to be an effective treatment, it would need to be administered as a long-acting therapy, but this might not be feasible due to the profound delay in gastric emptying induced by GLP-1. In order to assess the feasibility and efficacy of continuous administration of GLP-1 in NIDDM, we determined the effects of continuous intravenous infusion of GLP-1 (7-36) amide, from 22.00-17.00 hours, on glucose and insulin concentrations overnight and in response to three standard meals, in eight subjects with NIDDM. These were compared with responses to 0.9% NaCl infusion and responses in six non-diabetic control subjects who were not receiving GLP-1. Effects of beta-cell function were assessed in the basal state using homeostasis model assessment (HOMA) and in the postprandial state by dividing incremental insulin responses to breakfast by incremental glucose responses. To assess possible clinical benefit from priming of beta cells by GLP-1 given overnight only, a third study assessed the effect of GLP-1 given from 22.00-07.30 hours on subsequent glucose responses the next day. Continuous GLP-1 infusion markedly reduced overnight glucose concentrations (mean from 24.00-08.00 hours) from median (range) 7.8 (6.1-13.8) to 5.1 (4.0-9.2) mmol/l (p < 0.02), not significantly different from control subjects, 5.6 (5.0-5.8) mmol/l. Daytime glucose concentrations (mean from 08.00-17.00 hours) were reduced from 11.0 (9.3-16.4) to 7.6 (4.9-11.5) mmol/l (p < 0.02), not significantly different from control subjects, 6.7 (6.5-7.0) mmol/l. GLP-1 improved beta-cell function in the basal state from 62 (13-102) to 116 (46-180) %beta (p < 0.02) and following breakfast from 57 (19-185) to 113 (31-494) pmol/mmol (p < 0.02). GLP-1 only given overnight did not improve the glucose responses to meals the next day. In conclusion, continuous infusion of GLP-1 markedly reduced diurnal glucose concentrations, suggesting that continuous GLP-1 administration may be as useful therapy in NIDDM.

In situ localization of enzymes and mucin in normal rat colon embedded in plastic.

Several enzymes were investigated histochemically in the colons of normal male F344 rats in order to understand the function of different types of cells in this tissue. Serial methacrylate-embedded sections (2-4 microns) allowed the precise localizations of several enzymes including acid phosphatase, alkaline phosphatase, gamma-glutamyl transpeptidase, N-acetyl-beta-D-glucosaminidase (hexosaminidase), alpha-naphthyl butyrate esterase and 5'-nucleotidase. Sites reactive with periodic acid-Schiff were also localized. Gradients of enzyme activity were observed between caecum and rectum and/or from the luminal surfaces to the bases of the crypts for hexosaminidase, esterase and gamma-glutamyl transpeptidase. To our knowledge this is the first histochemical demonstration of gamma-glutamyl transpeptidase in normal rat colonic epithelial cells. The utilization of the methacrylate-embedding technique has revealed previously undescribed gradients of enzyme activity and has allowed the localization of enzyme activities not previously reported in normal rat colonic mucosa.

Tibial plateau fractures: evaluation with MR imaging.

Thirty patients with 31 tibial plateau fractures, initially diagnosed with standard radiography, were further studied with biplane linear tomography and magnetic resonance (MR) imaging. Each fracture was categorized according to the Schatzker classification. The extent of comminution and articular depression was determined for each modality. MR images were also evaluated for the presence of accompanying meniscal and ligamentous injury. In each case, MR imaging was noted to be as effective as tomography in depicting the amount of articular depression. MR imaging was found to be more effective than tomography in determining the extent of comminution. Beyond demonstrating the fracture, MR imaging was capable of revealing associated ligamentous and meniscal injuries. There was an increased prevalence of accompanying soft-tissue injuries in those fracture types that are associated with more violent forces and as the extent of articular depression increased. MR imaging is a useful and effective means of preoperatively evaluating tibial plateau fractures.

Prevalence of diabetes mellitus and impaired glucose tolerance in parents of women with gestational diabetes.

Nuclear families of non-insulin-dependent diabetic (NIDDM) patients are uncommon, as usually one or both parents have died. In order to aid identification of complete nuclear families, we have ascertained the disease process at a younger age by studying subjects with previous gestational diabetes. One hundred women who had had gestational diabetes, age (+/- SD) 38 (6) years, were screened by fasting plasma glucose (fpg). Sixty-one were found to have either fasting hyperglycaemia (5.5 < or = fpg < 7.8 mmol/l) or diabetes. Of these women 35 had both parents alive and the parents of 14 of these women agreed to the assessment of their metabolism by a continuous infusion of glucose with model assessment (CIGMA). Seven probands had impaired glucose tolerance (IGT) and seven were diabetic. They were age 35 (4) years and had body mass index (BMI) 26 (5) kg/m2. The parents were aged 62 (6) years and had BMI 29 (6) kg/m2 and their affection status was defined as presence of glucose intolerance (fpg or post-infusion achieved plasma glucose level > 2 SD of an age and obesity matched population). In the 14 families, five probands (36%) had neither parent affected, six (43%) had one parent affected and three (21%) had both parents affected. Only three probands had a parent with diabetes as defined by World Health Organisation criteria. We conclude that the study of women who have had gestational diabetes allows detection of probands with diabetes or impared glucose tolerance, who have both parents available for study.(ABSTRACT TRUNCATED AT 250 WORDS)

UKPDS 21: low prevalence of the mitochondrial transfer RNA gene (tRNA(Leu(UUR))) mutation at position 3243bp in UK Caucasian type 2 diabetic patients.

Some patients with Type 2 (non-insulin-dependent) diabetes mellitus possess a mitochondrial mutation in the tRNA(Leu(UUR)) gene at position 3243 bp. These subjects show a maternal mode of inheritance and often have hearing defects. In French and Japanese populations, this mutation may be present in 1-3% of subjects with a family history of diabetes. We assessed the prevalence of this mutation in newly diagnosed diabetic subjects in the UK white Caucasian population. The 3243 bp mutation was not detected in 500 randomly selected Type 2 diabetic subjects, 50 gestational diabetic subjects, and members of a MODY pedigree. Two of 748 (0.27%) Type 2 diabetic subjects with a family history of diabetes were found to possess the mutation. These subjects had an early age of diagnosis (M 38 years; F 36 years) and were non-obese. The male patient showed evidence of markedly impaired beta-cell function and deafness, while the female was not deaf, had approximately 50% of normal pancreatic function and responded well to diet. The mutation in the tRNA(Leu(UUR)) gene probably occurs in only approximately 0.1-0.2% of white Caucasian Type 2 diabetic patients in the UK.