Anti-inflammatory, procollagen, and wound repair properties of topical insulin gel

Diabetes mellitus is associated with impaired wound healing. The topical use of insulin is a promising therapy because it may favor all phases of the wound healing process. This study aimed to investigate the therapeutic outcomes of insulin gel in wounds of hyperglycemic mice. After diabetes induction, a 1-cm2 full-thickness wound was created on each animal's dorsum. The lesions were treated daily for 14 days with insulin gel (insulin group) or vehicle gel without insulin (vehicle group). Tissue samples were extracted on days 4, 7, 10, and 14 after the creation of the lesion. The samples were analyzed with hematoxylin/eosin and Sirius red staining, immunohistochemistry, Bio-Plex immunoassays, and western blotting. Insulin gel favored re-epithelialization at day 10 and increased the organization and deposition of collagen. Additionally, it modulated the expression of cytokines (interleukin (IL)-4 and IL-10) and increased the expression of arginase I, VEGF receptor 1, and VEGF on day 10. Activation of the insulin signaling pathway occurred via IRβ, IRS1, and IKK on day 10 and activation of Akt and IRS1 on day 14. These results suggested that insulin gel improved wound healing in hyperglycemic mice by modulating the expression of inflammatory factors, growth factors, and proteins of the insulin signaling pathway.

Metagenomics analysis reveals universal signatures of the intestinal microbiota in colorectal cancer, regardless of regional differences

The human gut microbiota is a complex and dynamic community of microorganisms living in our intestines and has emerged as an important factor for colorectal adenocarcinoma (CRC). The purpose of our study was to investigate the microbiota composition in Brazilian CRC patients compared with a local control population (CTL) to find out which changes could be considered universal or regional features in CRC microbiota. Fecal samples were obtained from 28 CRC and 23 CTL individuals. The 16S rRNA gene was used for metagenomic analysis. In addition to the anthropometric variables, the clinical stage (TNM 2018) was considered. Patients with CRC had a significant increase in alpha diversity and a higher percentage of genus Prevotella and a decreased proportion of Megamonas and Ruminococcus. Additionally, the proportion of Faecalibacterium prausnitzii was associated with a better prognosis in the first stages of CRC, and Fusobacterium nucleatum proved to be an important marker of colorectal carcinogenesis and tumor aggressiveness. Although regional differences influence the composition of the microbiota, in the case of CRC, the microhabitat created by the tumor seems to be a major factor. Our results contribute to a better understanding of the carcinogenic process, and even in different environments, some factors appear to be characteristic of the microbiota of patients with CRC.

Enhancement of cellular activity in hyperglycemic mice dermal wounds dressed with chitosan-alginate membranes

The use of specially designed wound dressings could be an important alternative to facilitate the healing process of wounds in the hyperglycemic state. Biocompatible dressings combining chitosan and alginate can speed up wound healing by modulating the inflammatory phase, stimulating fibroblast proliferation, and aiding in remodeling phases. However, this biomaterial has not yet been explored in chronic and acute lesions of diabetic patients. The aim of this study was to evaluate the effect of topical treatment with a chitosan-alginate membrane on acute skin wounds of hyperglycemic mice. Diabetes mellitus was induced by streptozotocin (60 mg · kg-1 · day-1 for 5 days, intraperitoneally) and the cutaneous wound was performed by removing the epidermis using a surgical punch. The results showed that after 10 days of treatment the chitosan and alginate membrane (CAM) group exhibited better organization of collagen fibers. High concentrations of interleukin (IL)-1α, IL-1β, granulocyte colony-stimulating factor (G-CSF), and tumor necrosis factor-alpha (TNF-α) were detected in the first and second days of treatment. G-CSF and TNF-α level decreased after 5 days, as well as the concentrations of TNF-α and IL-10 compared with the control group (CG). In this study, the inflammatory phase of cutaneous lesions of hyperglycemic mice was modulated by the use of CAM, mostly regarding the cytokines IL-1α, IL-1β, TNF-α, G-CSF, and IL-10, resulting in better collagen III deposition. However, further studies are needed to better understand the healing stages associated with CAM use.

A high-fructose diet induces insulin resistance but not blood pressure changes in normotensive rats

Rats fed a high-fructose diet represent an animal model for insulin resistance and hypertension. We recently showed that a high-fructose diet containing vegetable oil but a normal sodium/potassium ratio induced mild insulin resistance with decreased insulin receptor substrate-1 tyrosine phosphorylation in the liver and muscle of normal rats. In the present study, we examined the mean blood pressure, serum lipid levels and insulin sensitivity by estimating in vivo insulin activity using the 15-min intravenous insulin tolerance test (ITT, 0.5 ml of 6 æg insulin, iv) followed by calculation of the rate constant for plasma glucose disappearance (Kitt) in male Wistar-Hannover rats (110-130 g) randomly divided into four diet groups: control, 1:3 sodium/potassium ratio (R Na:K) diet (C 1:3 R Na:K); control, 1:1 sodium/potassium ratio diet (CNa 1:1 R Na:K); high-fructose, 1:3 sodium/potassium ratio diet (F 1:3 R Na:K), and high-fructose, 1:1 sodium/potassium ratio diet (FNa 1:1 R Na:K) for 28 days. The change in R Na:K for the control and high-fructose diets had no effect on insulin sensitivity measured by ITT. In contrast, the 1:1 R Na:K increased blood pressure in rats receiving the control and high-fructose diets from 117 + or - 3 and 118 + or - 3 mmHg to 141 + or - 4 and 132 + or - 4 mmHg (P<0.05), respectively. Triacylglycerol levels were higher in both groups treated with a high-fructose diet when compared to controls (C 1:3 R Na:K: 1.2 + or - 0.1 mmol/l vs F 1:3 R Na:K: 2.3 + or - 0.4 mmol/l and CNa 1:1 R Na:K: 1.2 + or - 0.2 mmol/l vs FNa 1:1 R Na:K: 2.6 + or - 0.4 mmol/l, P<0.05). These data suggest that fructose alone does not induce hyperinsulinemia or hypertension in rats fed a normal R Na:K diet, whereas an elevation of sodium in the diet may contribute to the elevated blood pressure in this animal model

A high-fructose diet induces changes in pp185 phosphorylation in muscle and liver of rats

Insulin stimulates the tyrosine kinase activity of its receptor resulting in the tyrosine phosphorylation of pp185, which contains insulin receptor substrates IRS-1 and IRS-2. These early steps in insulin action are essential for the metabolic effects of insulin. Feeding animals a high-fructose diet results in insulin resistance. However, the exact molecular mechanism underlying this effect is unknown. In the present study, we determined the levels and phosphorylation status of the insulin receptor and pp185 (IRS-1/2) in liver and muscle of rats submitted to a high-fructose diet evaluated by immunoblotting with specific antibodies. Feeding fructose (28 days) induced a discrete insulin resistance, as demonstrated by the insulin tolerance test. Plasma glucose and serum insulin and cholesterol levels of the two groups of rats, fructose-fed and control, were similar, whereas plasma triacylglycerol concentration was significantly increased in the rats submitted to the fructose diet (P<0.05). There were no changes in insulin receptor concentration in the liver or muscle of either group. However, insulin-stimulated receptor autophosphorylation was reduced to 72 + or - 4 percent (P<0.05) in the liver of high-fructose rats. The IRS-1 protein levels were similar in both liver and muscle of the two groups of rats. In contrast, there was a significant decrease in insulin-induced pp185 (IRS-1/2) phosphorylation, to 83 + or - 5 percent (P<0.05) in liver and to 77 + or - 4 percent (P<0.05) in muscle of the high-fructose rats. These data suggest that changes in the early steps of insulin signal transduction may have an important role in the insulin resistance induced by high-fructose feeding

Myocardial contractile response to an oral glucose load in normal subjects evaluated by echocardiography

There is a paucity of experimental data on the actual mechanism of insulin-induced changes on the myocardial function. In the present study we investigated the myocardial contractile, response to an oral glucose load using echocardiography. Fifteen healthy volunteers were studied after overnight fast and 150 minutes after the oral load of 75 g glucose. Oral glucose load caused an increase in plasma glucose and insulin levels, which was accompained by a significant increase in left ventricular shortening (from 35.2 + 0.7 per cent at baseline, to 38.5 + 0.6 per cent and 39 + 0.9 per cent at 30 and 60 minutes post glucose load, respectively [P<0.05 vs baseline]; ejection faction rose from 0.73 per cent + 0.01 to 0.77 per cent + 0.01 (P<0.05); pressure rate product increased from 7.29 + 0.2 to 8.31 + 0.3 mmHg x beats per min (P<0.007) and heart rate enhanced from 68.3 + 1.9 to 74 + 1.6 (P<0.034) and 75.3 + 1.5 beats per min (P<0.008) at 60 and 90 minutes after glucose, respectively. Meanwhile, mean arterial presure decreased significantly (10+1.5 per cent, P<0.018) when compared to basal values. These results indicate a significant change in the myocardial contractile response to an oral glucose load, probably related to baroreceptor reflex response as well as an overridden by a potent vasodilatador action of insulin. Nevertheless, we could not rule out the cardiac effects may also be due an insulin-induced sympathetic activation or a direct myocardial effect.

The insulin receptor substrate 1 associates with phosphotyrosine phosphatase SHPTP2 in liver and muscle of rats

Insulin stimulates the tyrosine kinase activity of its receptor resulting in the phosphorylation of its cytosolic substrate, insulin receptor substrate-1 (IRS-1) which, in turn, associates with proteins containing SH2 domains. It has been shown that IRS-1 associates with the tyrosine phosphatase SHPTP2 in cell cultures. While the effect of the IRS-1/SHPTP2 association on insulin signal transduction is not completely known, this association may dephosphorylate IRS-1 and may play a critical role in the mitogenic actions of insulin. However, there is no physiological demonstration of this pathway of insulin action in animal tissues. In the present study we investigated the ability of insulin to induce association between IRS-1 and SHPTP2 in liver and muscle of intact rats, by co-immunoprecipitation with anti-IRS-1 antibody and anti-SHPTP2 antibody. In both tissues there was an increase in IRS-1 association with SHPTP2 after insulin stimulation. This association occurred when IRS-1 had the highest level of tyrosine phosphorylation and the decrease in this association was more rapid than the decrease in IRS-1 phosphorylation levels. The data provide evidence against the participation of SHPTP2 in IRS-1 dephosphorylation in rat tissues, and suggest that the insulin signal transduction pathway in rat tissues is related mainly to the mitogenic effects of the hormone.

Insulin receptor has tyrosine kinase activity toward Shc in rat liver

Insulin induces tyrosine phosphorylation of Shc in cell cultures and in insulin-sensitive tissues of the intact rat. However, the ability of insulin receptor (IR) tyrosine kinase to phosphorylate Shc has not been previously demonstrated. In the present study, we investigated insulin-induced IR tyrosine kinase activity towards Shc. Insulin receptor was immunoprecipitated from liver extracts, before and after a very low dose of insulin into the portal vein, and incubated with immunopurified Shc from liver of untreated rats. The kinase assay was performed in vitro in the presence of exogenous ATP and the phosphorylation level was quantified by immunoblotting with antiphosphotyrosine antibody. The results demonstrate that Shc interacted with insulin receptor after infusion of insulin, and, more important, there was insulin receptor kinase activity towards immunopurified Shc. The description of this pathway in animal tissue may have an important role in insulin receptor tyrosine kinase activity toward mitogenic transduction pathways.

Viscosity of gums in vitro and their ability to reduce postprandial hyperglycemia in normal subjects

Experiments were carried out in vitro with three viscous polysaccharides (guar gum, pectin, and carboxymethylcellulose (CMC) of similar initial viscosity submitted to conditions that mimic events occurring in the stomach and duodenum, and their viscosity in these situations was compared to their actions on postprandial hyperglycemia in normal human subjects. Guar gum showed greater viscosity than the other gums during acidification and/or alkalinization and also showed larger effects on plasma glucose levels (35 per cent reduction in maximum rise in plasma glucose) and on the total area under the curve of plasma glucose (control: 20,314 + 1007 mg dl(-1) 180 min (-1) vs guar gum: 18,277 + 699 mg dl(-1) 180 min (-1), P<0.01). Pectin, which showed a marked reduction in viscosity at 37 degrees Celsius and after events mimicking those that occur in the stomach and duodenum, did not have a significant effect on postprandial hyperglycemia. The performance of viscosity and the glycemia response to CMC were at an intermediate level between guar gum and pectin. In conclusion, these data suggest that temperature, the process of acidification, alkalinization and exposure to intestinal ions induce different viscosity changes in gums having similar initial viscosity, establishing a direct relationship between a minor decrease of gum viscosity in vitro and a reduction of postprandial hyperglycemia.

Tissue-specific regulation of IRS-1 in unilaterally nephrectomized rats

Insulin stimulates the tyrosine kinase activity of its receptor, resulting in the phosphorylation of its cytosolic substrate, insulin receptor substrate 1 (IRS-1). IRS-1 is also a substrate for different peptides and growth factors, and a transgenic mouse "knockout" for this protein does not have normal growth. However, the role of IRS-1 in kidney hypertrophy and/or hyperplasia was not investigated. In the present study we investigated IRS-1 protein and tyrosine phosphorylation levels in the remnant kidney after unilateral nephrectomy (UNX) in 6-week-old male Wistar ratas. After insulin stimulation the levels of insulin receptor and IRS-1 tyrosine phosphorylation were reduced to 79 + 5 percent (P<0.005) and 58 + 6 percent (P<0.0001), respectively, of the control (C) levels, in the remnant kidney. It is possible that a circulating factor and/or a local (paracrine) factor playing a role in kidney growth can influence the early steps of insulin action in parallel. To investigate the hypothesis of a circulating factor, we studied the early steps of insulin action in liver and muscle of unilateral nephrectomized rats. There was no change in pp185 tyrosine phosphorylation levels in liver (C 100 + 12 percent vs UNX 89 + 9 percent, NS) and muscle (C 100 + 22 percent vs UNX 91 + 17 percent, NS), and also there was no change in IRS-1 phosphorylation levels in both tissues. These data demonstrate that after unilateral nephrectomy there is a decrease in insulin-induced insulin receptor and IRS-1 tyrosine phosphorylation levels in kidney but not in liver and muscle. It will be of interest to investigate which factors, probably paracrine ones, regulate these early steps of insulin action in the contralateral kidney of unilaterally nephrectomized rats.

Effect of fasting on insulin signaling in the aorta of intact rats

Insulin stimulates the tyrosine kinase activity of its receptor, resulting in the phosphorylation of its cytosolic substrate, insulin receptor substrate 1 (IRS-1). Previous studies have demonstrated a tissue-specific regulation of IRS-1. In the present study we investigated the levels and phosphorylation state of IRS-1 after insulin stimulation in the rat aorta in vivo, and the modulation of this protein after 72 h of fasting, using immunoprecipitation and immunoblotting with anti-insulin receptor, anti-IRS-1 and antiphosphotyrosine antibodies. We show that IRS-1 is present in rat aorta, and is tyrosine phosphorylated after insulin stimulation. After insulin stimulation, rats fasted for 72 h showed an increase in insulin receptor(l00 ñ 45 per cent, P<0.05)and IRS-1 phosphorylation (68 ñ 24 per cent, P<0.05) in aorta, compared to fed rats. There was no change in insulin receptor or IRS-1 protein levels in fasted rats. In summary, the present study demonstrated that proteins involved in the early steps of insulin signal transduction are present in the rat aorta and can be modulated by fasting. It will be of interest to study the regulation of these proteins in the aorta of animal models of hypertension and/or atherosclerosis.

Insulin resistance and myocardial hypertrophy in the attenuated reduction in mean arterial pressure after a glucose load in hypertensive patients

Although long recognized, the vasodilator effect of insulin has been relatively neglected over the last few years. Recent reports have focused on the sympathetic and antinatriuretic actions of this hormone. In the first part of the present study we characterized the reduction in blood pressure after a glucose load in hypertensive patients with and without insulin resistance. fourteen hypertensive Caucasian patients and ten Caucasian controls were submitted to a standard oral glucose tolerance test (OGTT) and intravenous insulin tolerance test (15-min ITT). In the hypertensive patients with insulin resistance the reduction in mean arterial presure (MAP) after a glucose load was blunted (6.7 ñ 1.7 per cent (N = 5)) when compared to insulin-sensitive (12.9 ñ 1.1 per cent (N = 9)) and normal subjects (10.1 ñ 0.8 per cent). IN the second part of the study we investigated whether hypertensive patients with myocardial hypertrophy were more insulin resistant than hypertensive individuals with a normal cardiac mass. The glucose disappearance rate (Kitt) was lower in hypertensive patients with myocardial myocardial hypertensive patients with myocardial hypertrophy (6.0 ñ 1.0 (N = 6)) when compared to hypertensive patients without myocardial hypertrophy (8.2 ñ 1.0 per cent/min (N = 8)), suggesting an association between this organomegaly and insulin resistance. In conclusion, our results suggested that 1) insulin resistance, rather than hyperinsulinemia, acts as a risk factor for the development of hypertension, because of insulin's inability to decrease MAP in this situation, and 2) there is an association between left ventricular hypertrophy and insulin resistance in hypertensive patients

Insulin resistance in psoriasis

The oral glucose tolerance test (OGTT) and intravenous insulin tolerance test (15-min ITT) were applied to ten patients with psoriasis and to 11 control subjects. No significant differences in mean plasma glucose levels were detected between psoriatic patients and normal individuals. In contrasts, serum insulin levels were significantly higher for the psoriatic patients as compared to the controls at 30, 60 and 120 min during the OGTT (P<0.05). The glucose disappearance rate during the 15-min ITT was lower in patients with psoriasis than in controls (5.1 + or - 0.5 percent min vs 7.5 + or - 0.4 percent/min, P<0.05), demonstrating a state of insulin resistance. Interestingly, the reduction in serum potassium levels during the ITT was also lower in the patients than in the controls (0.6 + or - 0.06 mEq/l vs 1.06 + or - 0.07 mEq/l,P<0.05), suggesting that the insulin resistance observed in psoriasis is not only related to glucose metabolism, but also to another important action of insulin, namely extrarenal potassium homeostasis

Molecular mechanisms of insulin resistance

1. Insulin stimulates tyrosine phosphorylation of the insulin receptor and of an endogenous substrate of approximately 185 kDa (insulin receptor substrate 1 or IRS-1). IRS-1 fulfills the criteria of a direct substrate of the insulin receptor, and tyrosine phosphorylation of IRS-1 leads to another step in insulin action, i.e., an association of phosphorylated IRS-1 with the enzyme PI3-kinase activating this enzyme. Using antipeptide antibodies to insulin receptor, to IRS-1 and to PI 3-kinase together with anti-phosphotyrosine antibodies it is possible to study insulin-stimulated insulin receptor phosphorylation, IRS-1 phosphorylation and the association/activation of IRS-1/PI 3-kinase. 2. In this review we describe alterations in these three early steps of insulin action after binding in animal models of insulin resistance, i.e., streptozotocin-induced diabetes (STZ diabetes), fasting, spontaneously hypertensive rats, the ob/ob mice, dexamethasone-treated rats, and the chronic effect of insulin on Fao cells in culture. 3. In states of insulin resistance with hypoinsulinemia (STZ diabetes and fasting) there is an increase in these early steps of insulin action. In animal models of insulin resistance with hyperinsulinemia there is a decrease in these steps of insulin action, indicating molecular post-receptor defects. Since we could reproduce the decrease in these three early steps of insulin action in cells in culture by chronic treatment with insulin, we postulate that these defects may be a consequence of the hyperinsulinemia of these animals.

Heterogenous insulin response to an oral glucose load by patients with the indeterminate clinical form of Chagas' disease

This study was designed to investigate the behavior of serum glucose and insulin in response to an oral glucose load in chagasic patients with the indeterminate clinical form of the disease. Sixteen chagasic patients and 28 healthy control subjects were studied after an overnight fast and during 2 h after ingestion of 100 g glucose. There were no significant differences in serum glucose levels before and 2 h after the glucose load between chagasic and control subjects. However, in 8 chagasic patients, the total area under the insulin curve was significantly lower (2976 +/- 448 microU ml-1 120 min-1) than in the control (10123 +/- 995 microU ml-1 120 min-1) and in the remaining chagasic patients (9220 +/- 826 microU ml-1 120 min-1). These results suggest that the hypoinsulinemia of this subgroup of chagasic patients may be secondary to reduced insulin secretion and/or to increased peripheral insulin sensitivity probably related to autonomic dysfunction

Endogenous hyperinsulinemia and release of growth hormone following a glucose load as well as in reponse to thyrotropin relasisng hormone in acromegaly

This study was designed to determine the relationship between endogenous hyperinsulinemia or free fatty acid (FFA) levels and the abnormal release of growth hormone (GH) during the oral glucose tolerance test (OGTT) or in response to thyrotropin releasing hormone (TRH) in acromegalic patients. Seventeen patients with acromegaly and nine healthy control subjects were studied. The 12 acromegalic patients who did not show the paradoxical response of increased GH secretion to the OGTT an/or to TRH exhibited higher insulin levels (total area under the curve during the OGTT) than patients who did. However, the FFA levels of the groups of acromegalic patients were similar. These data suggest that the endogenous hyperinsulinemia exhibited by many patients with acromegaly is associated with the absence of the paradoxical increase in GH secretion during the OGTT and/or after TRH

Association of specific histocompatibility antigens and acanthosis nigricans with insulin resistance

The association or interaction of histocompatibility antigens (HLA) and acanthosis nigricans with type A insulin resistance was studied in 13 patients (10 from family I, 2 from family II and an isolated case) for both sexes. HLA typing for the A, B, C and D antigens was performed by a standard microcytotoxicity test for all patients and for 100 normal controls from the same geographic area. The frequency of HLA B8 was 21% in the control group and 100% in patients with acanthosis nigricans. The frequency of HLA A1B8 was 15% in controls and 73% n acanthotic patients. All the members of family I presenting the association of a possibile insulin receptor defect (most likely provided by patient 2) with HLA B8 (provided by patient I) showed a more pronunced clinical and laboratory expression of insulin resistance. These data suggest that class I antigens of major histocompatibility complex (MHC), A1 and/or B8, may be involved in the pathogenesis of some forms of insulin resistance such as acanthosis nigricans (type A syndrome), possibly by a molecular interaction of the antigens with insulin receptors

Differences in first-phase insulin release between normal blacks and whites

This study was designed to compare insuslin release during the intravenous glucose tolerance test (GTT) in normal blacks and whites. The study included 14 normal blacks (9 males and 5 females) and 15 normal whites (10 males and 5 females). The index of first phase insulin release (sum of 1 + 3 min insulin levels and total area under the curve for the first 10 min of the GTT) was significantly higher in blacks. The difference is discussed in terms of genetic and acquired trats

Insulin resistance in essential hypertension

To determine the possible existence of a relationship between insulin resistance and sympthetic nervous system activity in essential hypertension, we calculated the double cross index for 14 hypertensive sujects and 14 normotensive subjects submitted to the oral glucose test. Plasma glucose and insulin levels were similar in hypertensive and normotensive subjects. After glucose loading, however, both parameters were significantly higher in hypertensive subjects. Five out of 14 hypertensive patients were hyperinsulinemic. The increase in double cross index following a glucose load was significantly higher in normotensive volunteers than in hyperinsulinemic hypertensive subjects. No change in double cross index was observed in normoinsulinemic hypertensive subjects. Thus, insulin resistance, high blood glucose level, impairment of cardiac response and hyperinsulinemia are present in a significant portion of hypertensive patients. Hyperinsulinemia may contribute to hypertensión by stimulating sympathetic nervous system activity, by influencing the calcium transport across the cell membrane and/or by some other mechanism

Peripheral muscle glucose and potassium transport in family with acanthosis nigricans and insulin resistance

This study was designed to determine the forearm exchange of glucose and potassium in four members of a family exhiting acanthosis nigricans and insulin resistance. Total areas under the curves for potassium release or uptake were within the normal range in all patients. Peripheral glucose uptake, however, was normal in 3 patients and lower in one patiente. In the latter patient, the dissociation observed between glucose and potassium transport suggests several manners of differential impairment of insulin action and/or steps distal to the insulin receptor as being responsible for insulin resistance