A proof of principle investigation of a novel enzyme formulation on dental calculus deposition: a 4-week randomized human clinical trial.

BACKGROUND: The dissolution of dental calculus, safely and at home, is among the more challenging issues facing the over-the-counter healthcare industry. Pontis Biologics, Inc. has developed novel model of calculus development and structure and has formulated a dentifrice (Tartarase™) using digestive enzymes as active ingredients that is shown to dissolve dental calculus in this Proof of Principle clinical trial. METHODS: This investigation was designed to evaluate the safety and efficacy of a novel enzyme formulation to remove existing calculus deposits in 4 weeks, measured using the Volpe-Manhold Index (V-MI) on lingual surfaces of 6 lower anterior teeth. The test formulation was compared to Crest Cavity Protection, as a control dentifrice. A total of 40 randomized test subjects began the study with 20 assigned to the control dentifrice and 20 assigned to the Tartarase groups (ten each, one brushing with Tartarase twice daily and one brushed with Tartarase and wore a dental tray filled with Tartarase for 30 min then brushed again with Tartarase, once daily). RESULTS: The Crest group experienced a 12% increase in calculus, in contrast to the results of both Tartarase groups that experienced a 40% reduction in calculus in 4 weeks of unsupervised at home use of the Tartarase toothpaste formulation. CONCLUSIONS: This proof of principle study demonstrates that a dentifrice, formulated along the lines of the Tartarase material, is capable of combating calculus accumulation using the same oral hygiene habits that are common worldwide. TRIAL REGISTRATION: This trial was registered retrospectively at clinicaltrials.gov and has the Unique Identification Number: NCT06139835, 14/11/2023.

Peripheral fat loss and decline in adipogenesis in older humans.

OBJECTIVE: Aging is associated with a redistribution of body fat including a relative loss of subcutaneous peripheral fat. These changes in body fat can have important clinical consequences since they are linked to increased risk of metabolic complications. The causes and mechanisms of loss of peripheral fat associated with aging are not clear. The aim of this study was to assess whether defects in adipogenesis contribute to fat loss in aging humans, as suggested from animal studies, and to evaluate the role of inflammation on pathogenesis of fat loss. MATERIALS/METHODS: Preadipocytes isolated from subcutaneous peripheral fat of healthy young and elderly subjects were compared in their ability to replicate and differentiate. RESULTS: The results show that both the rate of replication and differentiation of preadipocytes are reduced in older subjects. The reduction in adipogenesis is accompanied by a higher plasma level of the inflammatory marker, soluble tumor necrosis factor receptor 2, and greater release of tumor necrosis factor α from fat tissue. CONCLUSIONS: Thus, the gradual relative loss of peripheral fat in aging humans may in part result from a defect in adipogenesis, which may be linked to inflammation and increased release of proinflammatory cytokines from fat tissue.

Chromium Therapy for Insulin Resistance Associated with HIV-Disease.

OBJECTIVE: With the advent of highly active anti-retroviral therapy, HIV disease has become a chronic condition, but with a number of metabolic complications including insulin resistance and diabetes mellitus, dyslipidemia and hypertension and an increased incidence of atherosclerosis. The aim of the current study was to test the safety and efficacy of chromium picolinate for HIV- associated insulin resistance. MATERIALS/METHODS: The study was a randomized, double-blind, placebo-controlled trial with subjects receiving 500µg of chromium picolinate or placebo twice daily for two months. HIV- infected subjects were selected based on a fasting concentration of plasma glucose greater than 5.5mmol/L or a plasma glucose concentration of greater than 7.7mmol/L (but less than 11mmol/L) 2h after oral ingestion of 75g of glucose. Insulin sensitivity was assessed with a hyper-insulinemic-euglycemic clamp and glucose tolerance was assessed with the oral glucose tolerance test. Subjects were monitored closely for alterations in viral load, CD4+ cells, hemoglobin and hematocrit, kidney and liver function, and fasting lipid profiles. RESULTS: Forty-three subjects were enrolled and 39 completed the protocol (20 in the chromium-supplemented and 19 in the placebo arm). Following chromium-supplementation, there were no significant changes in either insulin sensitivity or glucose tolerance. There was a significant improvement in serum HDL cholesterol concentration in the group supplemented with chromium. CONCLUSIONS: Chromium picolinate supplementation at this level was well-tolerated, but overall was not an effective therapy for insulin resistance in these HIV-infected subjects.

Rosiglitazone-Mediated Effects on Skeletal Muscle Gene Expression Correlate with Improvements in Insulin Sensitivity in Individuals with HIV-Insulin Resistance.

Rosiglitazone, an agonist of peroxisome proliferator activated receptor (PPARγ), improves insulin sensitivity by increasing insulin-stimulated glucose uptake into muscle tissue. This study was undertaken to assess changes in expression of PPAR-regulated genes in muscle tissue following treatment of HIV-associated insulin resistance with rosiglitazone. Muscle gene expression was assessed in twenty-two seronegative HIV subjects (control), 21 HIV-infected individuals with normal insulin sensitivity (HIV-IS) and 19 HIV-infected individuals with insulin resistance (HIV-IR). A subset of the HIV-IR group (N = 10) were re-evaluated 12 weeks after treatment with 8 mg/d of rosiglitazone. The HIV-IR group's rosiglitazone-mediated improvement in insulin sensitivity was highly correlated with increased expression of PPARγ and carnitine palmitoyl transferase-1 (CPT-1), (r = 0.87, P < .001) and (r = 0.95, P < .001), respectively. The changes in PPARγ expression were also correlated with the changes in CPT1 expression (r = 0.75, P = .009). The results suggest that rosiglitazone; may have a direct effect on muscle tissue to improve insulin sensitivity.

Delivery of rosiglitazone from an injectable triple interpenetrating network hydrogel composed of naturally derived materials.

An in situ gelable and biodegradable triple-interpenetrating network (3XN) hydrogel, completely devoid of potentially cytotoxic extraneous small molecule crosslinkers, is formulated from partially oxidized dextran (Odex), teleostean and N-carboxyethyl chitosan (CEC). Both the rheological profile and mechanical strength of the 3XN hydrogel approximate the combined characteristics of the three individual hydrogels composed of the binary partial formulations (i.e., Odex/CEC, Odex/teleostean, and CEC/teleostean). The 3XN hydrogel is considerably more resistant to fibroblast-mediated degradation compared to each partial formulation in cell culture models; this is attributable to the interpenetrating triple-network structure. The presence of teleostean in the 3XN hydrogel imparts cell affinity, constituting an environment amenable to fibroblast growth. in vivo subdermal injection into mouse model shows that the 3XN hydrogel does not induce extensive inflammatory response nor is there any evidence of tissue necrosis, further confirming the non-cytotoxicity of the hydrogel and its degradation byproducts. Importantly, the capability of the 3XN hydrogel to serve as a sustained drug delivery vehicle is confirmed using rosiglitazone as a model drug. The presence of rosiglitazone profoundly changes the cell/tissue interactions with the subdermally injected 3XN hydrogel. Rosiglitazone suppresses both the inflammatory response and tissue repair in a dose-dependent manner and considerably moderated the hydrogel degradation.

Effect of ritonavir and atazanavir on human subcutaneous preadipocyte proliferation and differentiation.

Protease inhibitors (PIs) have been implicated in the development of HIV-associated lipodystrophy through a reduction in the differentiation of preadipocytes. While atazanavir (ATV) is associated with fewer clinical metabolic abnormalities in the short-term, the effects of long-term exposure are not known. ATV effects on preadipocyte replication or differentiation would indicate the potential for long-term problems. This study compared ritonavir (RTV) and ATV effects on preadipocyte replication and differentiation in human primary cultures. Preadipocytes from subcutaneous fat were studied in the presence of therapeutic concentrations of RTV and ATV for replication, differentiation, and adipokine secretion. The effects of the drugs on the expression of PPARgamma and related genes during differentiation were also assessed by real-time quantitative PCR. RTV induced a significant inhibition of preadipocyte proliferation, differentiation and adiponectin secretion. ATV at concentrations within the range of therapeutic levels did not affect differentiation or adiponectin secretion, but did have inhibitory effects on preadipocyte proliferation. Inhibition of differentiation by PIs was associated with decreased expression of PPARgamma, C/EBPalpha, and aP2 genes. In summary, although ATV at therapeutic levels has a smaller impact on adipogenesis, alterations in preadipocyte proliferation suggest the potential for adverse effects with long-term use.

Importance of the high-molecular-mass isoform of adiponectin in improved insulin sensitivity with rosiglitazone treatment in HIV disease.

The present study was designed to investigate the relationship of isoforms of adiponectin to insulin sensitivity in subjects with HIV-associated insulin resistance in response to treatment with the thiazolidinedione, rosiglitazone. The two isoforms of adiponectin, HMW (high-molecular-mass) and LMW (low-molecular-mass), were separated by sucrose-gradient-density centrifugation. The amount of adiponectin in gradient fractions was determined by ELISA. Peripheral insulin sensitivity (Rd) was determined with hyperinsulinaemic-euglycaemic clamp, whereas hepatic sensitivity [HOMA (Homoeostasis Model Assessment) %S] was based on basal glucose and insulin values. Treatment with rosiglitazone for 3 months resulted in a significant improvement in the index of hepatic insulin sensitivity (86.4+/-15% compared with 139+/-23; P=0.007) as well as peripheral insulin sensitivity (4.04+/-0.23 compared with 6.17+/-0.66 mg of glucose/kg of lean body mass per min; P<0.001). Improvement in HOMA was associated with increased levels of HMW adiponectin (r=0.541, P=0.045), but not LMW adiponectin. The present study suggests that the HMW isoform of adiponectin is important in the regulation of rosiglitazone-mediated improvement in insulin sensitivity in individuals with HIV-associated insulin resistance, particularly in the liver.

Influence of age on the association of retinol-binding protein 4 with metabolic syndrome.

OBJECTIVE: The relationships of retinol-binding protein 4 (RBP4) with insulin sensitivity and body fat distribution have been investigated in a few recent studies with conflicting results. This may have been due to differences in ages of the subjects in the different studies. The aim of this study was to investigate whether the association of RBP4 and insulin sensitivity and percent trunk fat are influenced by age. METHODS AND PROCEDURES: Cross-sectional analyses of 48 young subjects and 55 elderly subjects. Insulin sensitivity was determined by a hyperinsulinemic-euglycemic clamp. Body fat distribution was determined by a dual-energy X-ray absorptiometry (DXA). RESULTS: In the young subjects, RBP4 levels were associated with insulin sensitivity (r = -0.30, P = 0.04), percent trunk fat (r = 0.54, P < 0.001), triglycerides (r = 0.44, P = 0.003), low-density lipoprotein (r = 0.38, P = 0.01). In contrast, in the elderly subjects there was no correlation between RBP4 levels and insulin sensitivity (r = -0.18, P = 0.20), percent trunk fat (r = 0.00, P = 0.10), triglycerides (r = 0.25, P = 0.10), and low-density lipoprotein (r = -0.11, P = 0.47). DISCUSSION: The associations of RBP4 with insulin sensitivity, percent trunk fat, and lipid levels are influenced by age.

Endothelial adhesion molecules are associated with inflammation in subjects with HIV disease.

BACKGROUND: The presence of the adhesion molecules intercellular adhesion molecule-1 (ICAM-1) and soluble vascular cell adhesion molecule-1 (VCAM-1) is associated with elevated risk of cardiovascular disease. Subjects with human immunodeficiency virus (HIV) disease have multiple risk factors for cardiovascular disease, including elevated serum lipid levels, insulin resistance, and elevated levels of ICAM-1 and VCAM-1. This study assessed the variables associated with elevated adhesion molecule levels in this patient population. METHODS: Serum levels of ICAM-1 and VCAM-1 were assessed in 31 subjects without HIV disease and 52 subjects with HIV disease. Pearson correlation indicated a significant relationship between ICAM concentration and other variables, including CD4+ cell count, HIV viral burden, insulin sensitivity, and serum lipid level. Multiple regression modeling was used to determine the strengths of association among the variables. RESULTS: Subjects with HIV disease had elevated levels of ICAM-1 and VCAM-1. Pearson correlation analysis revealed significant associations between ICAM-1 and VCAM-1 level and insulin sensitivity, plasma lipid level, and presence of type 2 soluble receptor for tumor necrosis factor-alpha (sTNFR2). With multiple regression modeling to control for interdependence, only sTNFR2, a marker of inflammation, was an independent predictor of ICAM-1 and VCAM-1 levels. CONCLUSIONS: The study suggests that many of the variables associated with ICAM-1 and VCAM-1 levels can be related to their impact on inflammation.

Limb fat to trunk fat ratio in elderly persons is a strong determinant of insulin resistance and adiponectin levels.

BACKGROUND: Similar to lipodystrophy syndromes, aging results in increased visceral adiposity with loss of subcutaneous adipose tissue in the extremities. The hypothesis of this study is that the distribution of limb fat to trunk fat (LF/TF) ratio in elderly persons has a stronger correlation than trunk fat alone to insulin resistance and adiponectin levels. METHODS: Thirty-eight elderly participants were divided into an insulin-resistant (IR) group and an insulin-sensitive (IS) group. Limb fat and trunk fat were measured by dual-energy x-ray absorptiometry. Insulin resistance was measured by a hyperinsulinemic-euglycemic clamp. RESULTS: There was no significant difference between the IS and IR groups with respect to body mass index, body fat index, absolute amount of trunk fat, or percent body fat. However, the difference in LF/TF ratio between the IS (1.02 +/- 0.05) and the IR groups (0.77 +/- 0.05) was highly significantly different (p <.001). Insulin resistance had a stronger correlation to the LF/TF ratio (r = 0.61, p <.001) than to absolute trunk fat (r = -0.32, p =.051). Adiponectin levels had a strong association with the LF/TF ratio (r = 0.63, p <.001), but did not correlate to absolute trunk fat (r = -0.24, p =.18). CONCLUSIONS: The distribution of body fat (LF/TF ratio) in elderly persons is a stronger determinant of insulin resistance and adiponectin levels than is trunk fat alone. The LF/TF ratio can be a useful tool to assess insulin sensitivity in the elderly population.

Wall suction applied to needle muscle biopsy - a novel technique for increasing sample size.

BACKGROUND: The needle biopsy technique described by Bergström is the most commonly used technique to obtain samples to assess muscle metabolism. Sampling of muscle, particularly the vastus lateralis, has become an essential tool in biomedical and clinical research. Optimal sample size is critical for availability of tissue for processing. To evaluate the effectiveness of a novel technique to obtain adequate sample size using wall suction applied to needle muscle biopsy, we collected samples from subjects in on-going clinical studies for gene expression. MATERIALS AND METHODS: Muscle biopsy samples of the vastus lateralis using 6 mm Bergström needles under local anesthesia were obtained from 55 subjects who had volunteered to participate in this research project. The vastus lateralis was biopsied according to the methods described by Bergström with a 6 mm biopsy needle. Wall suction was applied to the inner bore of the biopsy needle after the needle was inserted into the muscle. RESULTS: The mean sample of biopsy taken using the 6 mm was 233 mg (n = 55). The wall suction (200 mm Hg) applied to the needle pulled the surrounding tissue into the central bore of the needle. The quality of the samples was adequate for all biochemical assays. The biopsy technique did not result in any complications due to infection or bleeding. CONCLUSIONS: Using a novel technique of connecting a 6 mm Bergström biopsy needle to wall suction, we have obtained 200 to 300 mg muscle biopsy specimens uniformly, with ease, and minimal discomfort. An increase in sample size allows for a wider variety of biochemical and histopathological analysis.

Retinol-binding protein 4 is associated with insulin resistance and body fat distribution in nonobese subjects without type 2 diabetes.

BACKGROUND: Adipose tissue is responsible for releasing various adipokines that have been related to insulin resistance. Understanding the relationship of these adipokines to insulin resistance may foster the development of new treatments for diabetes. OBJECTIVES: The primary objective of this study was to determine whether an association between retinol-binding protein 4 (RBP4) and insulin resistance exists in nonobese individuals without a family history or diagnosis of diabetes. The secondary objective was to determine by a dual energy x-ray absorptiometry scan which adipose tissue depot most closely relates to RBP4 levels. DESIGN: Cross-sectional analysis of 92 study participants ranging in age from 20 to 83 yr was performed. The range of body mass index (BMI) was from 18 to 30 kg/m(2). Exclusion criteria were a BMI greater than 30 kg/m(2), family history of diabetes, or a diagnosis of diabetes. Insulin sensitivity was determined by a hyperinsulinemic euglycemic clamp. Body fat was measured by dual energy x-ray absorptiometry scan. RESULTS: RBP4 values were lower in females (35.8 +/- 1.7 microg/ml) compared with males (39.9 +/- 1.4 microg/ml; P = 0.06). RBP4 levels were found to correlate negatively with insulin sensitivity (r = -0.32; P = 0.002) and positively with age (r = 0.38; P < 0.001). RBP4 levels did not correlate with BMI (r = -0.13; P = 0.22), trunk fat (r = 0.16; P = 0.22), or percent body fat (r = 0.07; P = 0.65). However, RBP4 levels did correlate with percent trunk fat (r = 0.36; P = 0.001). CONCLUSION: These findings indicate a relationship between RBP4, insulin sensitivity, and percent trunk fat in individuals who may not have features of insulin resistance.

Chronic loss of subcutaneous adipose tissue in HIV-associated lipodystrophy may not be associated with accelerated apoptosis.

HIV-associated lipodystrophy (HIV-LD) is characterized by a loss of adipose tissue from the subcutaneous compartment. Previously reported data suggested that this loss of adipose tissue was the result of an increased rate of apoptosis in subcutaneous adipose tissue (SAT). The present study examined the rate of apoptosis in SAT with a sensitive ligase-mediated polymerase chain reaction technique to amplify DNA ladders. Individuals with HIV-LD were compared with HIV-infected subjects without LD and subjects without HIV disease. Although apoptosis was observed in subjects with HIV-LD, there was no difference in the incidence of individuals with apoptosis among those with HIV-LD (10 of 22 subjects), those with HIV but no LD (13 of 25 subjects), and those without HIV disease (13 of 27 subjects). These data suggest that HIV-related chronic loss of SAT may not always be associated with increased frequency of adipocyte apoptosis.

Loss of subcutaneous adipose tissue in HIV-associated lipodystrophy is not due to accelerated apoptosis.

HIV-associated lipodystrophy is characterized by a loss of adipose tissue from the subcutaneous compartment. Previously reported data suggested that this loss of adipose tissue was the result of an increased rate of apoptosis in subcutaneous adipose tissue. The present study examined the rate of apoptosis in subcutaneous adipose tissue with a sensitive ligase-mediated polymerase chain reaction technique to amplify DNA ladders. Individuals with HIV lipodystrophy were compared with HIV-infected subjects without lipodystrophy and subjects without HIV disease. Although apoptosis was observed in subjects with HIV lipodystrophy, there was no difference in the frequency of individuals with apoptosis among those with HIV lipodystrophy (10/22), those with HIV but no lipodystrophy (13/25), and subjects without HIV disease (13/27).

Adiponectin and leptin levels in HIV-infected subjects with insulin resistance and body fat redistribution.

In this study, we sought to determine the relationship between serum levels of leptin and adiponectin (Acrp30) in patients with HIV-associated lipodystrophy (HIV-LD). Three groups of subjects were studied; HIV-positive subjects with lipodystrophy (HIV-LD; n = 22), HIV-positive subjects without lipodystrophy (HIV; n = 17), and ethnicity- and body mass index-matched healthy control subjects (n = 20). Although total body fat from dual energy x-ray absorptiometry was similar in all three groups, the HIV-LD group had a significantly lower mean proportion of body fat in the limbs +/- SEM (37.2% +/- 2.2%) than either controls (49.8% +/- 1.5%) or HIV subjects (45.7% +/- 2.0%). The HIV-LD group also had the lowest mean insulin sensitivity +/- SEM (5.11 +/- 0.59 mg of glucose/[kg of lean body mass. min] vs. 10.2 +/- 0.72 mg of glucose/[kg of lean body mass. min] in controls and 8.64 +/- 0.69 mg of glucose/[kg of lean body mass. min] in the HIV group). Leptin levels were similar in all three groups and were significantly correlated to total body fat (r = 0.86; p <.001), but these levels did not correlate with either insulin sensitivity or limb fat. Mean Acrp30 levels +/- SEM were lowest in the HIV-LD group (5.43 +/- 0.44 microg/mL vs. 11.2 +/- 1.4 microg/mL in the HIV group and 14.9 +/- 1.8 microg/mL in control subjects). Further, Acrp30 levels were positively correlated with insulin sensitivity (r = 0.610; p <.001) and limb fat (r = 0.483; p <.001). However, the correlation between limb fat and insulin sensitivity disappeared when Acrp30 level and other potential mediators were removed from the association, suggesting that a deficiency in Acrp30 in subjects with HIV-LD may be part of the mechanism for the reduced insulin sensitivity.

Improved insulin sensitivity and body fat distribution in HIV-infected patients treated with rosiglitazone: a pilot study.

The insulin-sensitizing drugs thiazolidinediones (TZDs), such as rosiglitazone, improve insulin sensitivity and also promote adipocyte differentiation in vitro. The authors hypothesized that TZDs might be beneficial to patients with HIV disease to improve insulin sensitivity and the distribution of body fat by increasing peripheral fat. The ability of rosiglitazone (8 mg/d) to improve insulin sensitivity (from hyperinsulinemic-euglycemic clamp) and to improve body fat distribution (determined from computed tomography measurements of visceral adipose tissue [VAT] and subcutaneous adipose tissue [SAT]) was determined in 8 HIV-positive patients. Before treatment, the insulin sensitivity of the patients was reduced to approximately 34% of that in control subjects. The rate of glucose disposal during a hyperinsulinemic-euglycemic clamp (Rd) was 3.8 +/-.4 (SEM) mg glucose/kg lean body mass/min compared with 11.08 +/- 1.1 (p<.001) in healthy age- and body mass index (BMI)-matched control subjects. After rosiglitazone treatment of 6 to 12 weeks, Rd increased to 5.99 +/-.9 (p=.02), an improvement of 59 +/- 22%. SAT increased by 23 +/- 10% (p=.05), and, surprisingly, VAT was decreased by 21 +/- 8% (p=.04) with a trend for increased SAT/VAT that failed to reach statistical significance. There were no significant changes in blood counts, viral loads, or CD4 counts with rosiglitazone treatment. The study demonstrates that rosiglitazone therapy improves insulin resistance and body fat distribution in some patients with HIV disease.

Soluble tumour necrosis factor alpha receptor 2, a serum marker of resistance to the anabolic actions of growth hormone in subjects with HIV disease.

Therapies are still being sought for the prevention of loss of body weight and lean body mass in HIV disease. The purpose of the present study was to identify a serum marker that would help in selecting patients who may be appropriate candidates for the use of anabolic agents, such as growth hormone, to restore lean body mass. This study included 26 HIV-infected patients and nine healthy controls, assessed previously for the effectiveness of 2 weeks of growth hormone administration in the stimulation of protein synthesis in skeletal muscle. Serum levels of interleukins-1beta, -6 and -10 were not useful predictors of the anabolic response to growth hormone. Serum concentrations of tumour necrosis factor alpha (TNFalpha) were significantly elevated (P<0.05) in patients with AIDS and AIDS-related weight loss, and there was a significant correlation between the serum concentration of interleukin-1 receptor antagonist and stage of disease (P=0.03). However, the serum concentration of the soluble TNFalpha receptor type 2 was most predictive of an inability of muscle protein synthesis to respond anabolically to growth hormone (r=-0.42, P=0.01). These data suggest that inflammation impacts on the responsiveness of muscle tissue to an anabolic stimulus, and that the soluble TNFalpha receptor type 2 provides a useful serum marker for metabolic dysfunction in HIV disease, which can be used to identify individuals likely to respond to growth hormone-based anabolic therapy.