Aaron J. Kowalski

Continuous glucose monitoring and intensive treatment of type 1 diabetes

BACKGROUND The value of continuous glucose monitoring in the management of type 1 diabetes mellitus has not been determined. METHODS In a multicenter clinical trial, we randomly assigned 322 adults and children who were already receiving intensive therapy for type 1 diabetes to a group with continuous glucose monitoring or to a control group performing home monitoring with a blood glucose meter. All the patients were stratified into three groups according to age and had a glycated hemoglobin level of 7.0 to 10.0%. The primary outcome was the change in the glycated hemoglobin level at 26 weeks. RESULTS The changes in glycated hemoglobin levels in the two study groups varied markedly according to age group (P=0.003), with a significant difference among patients 25 years of age or older that favored the continuous-monitoring group (mean difference in change, -0.53%; 95% confidence interval [CI], -0.71 to -0.35; P<0.001). The between-group difference was not significant among those who were 15 to 24 years of age (mean difference, 0.08; 95% CI, -0.17 to 0.33; P=0.52) or among those who were 8 to 14 years of age (mean difference, -0.13; 95% CI, -0.38 to 0.11; P=0.29). Secondary glycated hemoglobin outcomes were better in the continuous-monitoring group than in the control group among the oldest and youngest patients but not among those who were 15 to 24 years of age. The use of continuous glucose monitoring averaged 6.0 or more days per week for 83% of patients 25 years of age or older, 30% of those 15 to 24 years of age, and 50% of those 8 to 14 years of age. The rate of severe hypoglycemia was low and did not differ between the two study groups; however, the trial was not powered to detect such a difference. CONCLUSIONS Continuous glucose monitoring can be associated with improved glycemic control in adults with type 1 diabetes. Further work is needed to identify barriers to effectiveness of continuous monitoring in children and adolescents. (ClinicalTrials.gov number, NCT00406133.)

Mice lacking osteopontin show normal development and bone structure but display altered osteoclast formation in vitro.

We have used homologous recombination in embryonic stem cells to generate mice with a targeted disruption of the osteopontin (Opn, or Spp1, for secreted phosphoprotein 1) gene. Mice homozygous for this disruption fail to express osteopontin (OPN) as assessed at both the mRNA and protein level, although an N‐terminal fragment of OPN is detectable at extremely low levels in the bones of −/− animals. The Opn−/− mice are fertile, their litter size is normal, and they develop normally. The bones and teeth of animals not expressing OPN are morphologically normal at the level of light and electron microscopy, and the skeletal structure of young animals is normal as assessed by radiography. Ultrastructurally, proteinaceous structures normally rich in OPN, such as cement lines, persist in the bones of the Opn−/− animals. Osteoclastogenesis was assessed in vitro in cocultures with a feeder layer of calvarial osteoblast cells from wild‐type mice. Spleen cells from Opn−/− mice cells formed osteoclasts 3‐ to 13‐fold more frequently than did control Opn+/+ cells, while the extent of osteoclast development from Opn−/− bone marrow cells was about 2‐ to 4‐fold more than from the corresponding wild‐type cells. Osteoclast development occurred when Opn−/− spleen cells were differentiated in the presence of Opn−/− osteoblasts, indicating that endogenous OPN is not required for this process. These results suggest that OPN is not essential for normal mouse development and osteogenesis, but can modulate osteoclast differentiation.

The effect of continuous glucose monitoring in well-controlled type 1 diabetes.

OBJECTIVE The potential benefits of continuous glucose monitoring (CGM) in the management of adults and children with well-controlled type 1 diabetes have not been examined. RESEARCH DESIGN AND METHODS A total of 129 adults and children with intensively treated type 1 diabetes (age range 8–69 years) and A1C <7.0% were randomly assigned to either continuous or standard glucose monitoring for 26 weeks. The main study outcomes were time with glucose level ≤70 mg/dl, A1C level, and severe hypoglycemic events. RESULTS At 26 weeks, biochemical hypoglycemia (≤70 mg/dl) was less frequent in the CGM group than in the control group (median 54 vs. 91 min/day), but the difference was not statistically significant (P = 0.16). Median time with a glucose level ≤60 mg/dl was 18 versus 35 min/day, respectively (P = 0.05). Time out of range (≤70 or >180 mg/dl) was significantly lower in the CGM group than in the control group (377 vs. 491 min/day, P = 0.003). There was a significant treatment group difference favoring the CGM group in mean A1C at 26 weeks adjusted for baseline (P < 0.001). One or more severe hypoglycemic events occurred in 10 and 11% of the two groups, respectively (P = 1.0). Four outcome measures combining A1C and hypoglycemia data favored the CGM group in comparison with the control group (P < 0.001, 0.007, 0.005, and 0.003). CONCLUSIONS Most outcomes, including those combining A1C and hypoglycemia, favored the CGM group. The weight of evidence suggests that CGM is beneficial for individuals with type 1 diabetes who have already achieved excellent control with A1C <7.0%.

International Consensus on Use of Continuous Glucose Monitoring

Measurement of glycated hemoglobin (HbA1c) has been the traditional method for assessing glycemic control. However, it does not reflect intra- and interday glycemic excursions that may lead to acute events (such as hypoglycemia) or postprandial hyperglycemia, which have been linked to both microvascular and macrovascular complications. Continuous glucose monitoring (CGM), either from real-time use (rtCGM) or intermittently viewed (iCGM), addresses many of the limitations inherent in HbA1c testing and self-monitoring of blood glucose. Although both provide the means to move beyond the HbA1c measurement as the sole marker of glycemic control, standardized metrics for analyzing CGM data are lacking. Moreover, clear criteria for matching people with diabetes to the most appropriate glucose monitoring methodologies, as well as standardized advice about how best to use the new information they provide, have yet to be established. In February 2017, the Advanced Technologies & Treatments for Diabetes (ATTD) Congress convened an international panel of physicians, researchers, and individuals with diabetes who are expert in CGM technologies to address these issues. This article summarizes the ATTD consensus recommendations and represents the current understanding of how CGM results can affect outcomes.

Exercise management in type 1 diabetes: A consensus statement

Type 1 diabetes is a challenging condition to manage for various physiological and behavioural reasons. Regular exercise is important, but management of different forms of physical activity is particularly difficult for both the individual with type 1 diabetes and the health-care provider. People with type 1 diabetes tend to be at least as inactive as the general population, with a large percentage of individuals not maintaining a healthy body mass nor achieving the minimum amount of moderate to vigorous aerobic activity per week. Regular exercise can improve health and wellbeing, and can help individuals to achieve their target lipid profile, body composition, and fitness and glycaemic goals. However, several additional barriers to exercise can exist for a person with diabetes, including fear of hypoglycaemia, loss of glycaemic control, and inadequate knowledge around exercise management. This Review provides an up-to-date consensus on exercise management for individuals with type 1 diabetes who exercise regularly, including glucose targets for safe and effective exercise, and nutritional and insulin dose adjustments to protect against exercise-related glucose excursions.

Osteopontin : An intrinsic inhibitor of inflammation in cartilage

OBJECTIVE To identify extracellular and intraarticular matrix components that are differentially expressed in normal and osteoarthritis (OA)-affected cartilage and to investigate their functions with respect to regulation of mediators of inflammation. METHODS Differential-display reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of a pool of messenger RNA (mRNA) from 10 human OA cartilage samples and 5 normal cartilage samples was performed using arbitrary primers. Confirmatory analysis of the up-regulated transcripts of fibronectin (FN) and osteopontin (OPN) was performed by RT-PCR of individual RNA samples from a separate set of donors. The effect of recombinant OPN (or anti-OPN antiserum) on chondrocyte function was examined by analyzing the spontaneous or interleukin-1 (IL-1)-induced release of nitric oxide (NO) and prostaglandin E2 (PGE2) from human OA-affected cartilage under ex vivo conditions. RESULTS Up-regulation (300-700%) of FN and OPN mRNA was observed in human OA-affected cartilage as compared with normal cartilage. Functional analysis of the role of OPN in OA cartilage showed that 1) Addition of 1 microg/ml (20 nM) of recombinant OPN to human OA-affected cartilage under ex vivo conditions inhibited spontaneous and IL-1beta-induced NO and PGE2 production, and 2) neutralization of intraarticular OPN with anti-OPN antiserum augmented NO production. CONCLUSION The data indicate that one of the functions of intraarticular OPN, which is overexpressed in OA cartilage, is to act as an innate inhibitor of IL-1, NO, and PGE2 production. These findings suggest that the production of pleiotropic mediators of inflammation that influence cartilage homeostasis, such as NO and PGE2, is regulated by the interaction of chondrocytes with differentially expressed proteins within the extracellular matrix.

Pathway to Artificial Pancreas Systems Revisited: Moving Downstream

Artificial pancreas (AP) systems, a long-sought quest to replicate mechanically islet physiology that is lost in diabetes, are reaching the clinic, and the potential of automating insulin delivery is about to be realized. Significant progress has been made, and the safety and feasibility of AP systems have been demonstrated in the clinical research center and more recently in outpatient “real-world” environments. An iterative road map to AP system development has guided AP research since 2009, but progress in the field indicates that it needs updating. While it is now clear that AP systems are technically feasible, it remains much less certain that they will be widely adopted by clinicians and patients. Ultimately, the true success of AP systems will be defined by successful integration into the diabetes health care system and by the ultimate metric: improved diabetes outcomes.

Characterization of anti-osteopontin monoclonal antibodies: Binding sensitivity to post-translational modifications†

Osteopontin (OPN) is primarily a secreted phosphoglycoprotein found in a variety of tissues and body fluids. It has a wide range of reported functions, many of which are affected by the degree of post‐translational modification (PTM) of the protein. These PTMs include phosphorylation, glycosylation, and cross‐linking by transglutaminase. Here we describe the generation of unique monoclonal antibodies raised against recombinant OPN utilizing the OPN knockout mouse. The antibodies exhibit differential binding to OPN produced by different cell lines from the same species, as well to the multiple OPN forms in human urine. Most of the antibodies generated are able to recognize OPN produced by ras‐transformed mouse fibroblasts, however only one antibody recognizes the more phosphorylated protein produced by the differentiating pre‐osteoblast murine cell line MC3T3E1. Using a novel biopanning procedure combining T7 phage gene fragment display and protein G precipitation, we have epitope‐mapped these antibodies. Several of the antibodies bind to regions of the OPN molecule that are phosphorylated, and one binds the region of OPN that is glycosylated. Using phosphorylated and non‐phosphorylated peptides, we show that the binding of two antibodies to the C‐terminal end of OPN is inhibited by phosphorylation of this region. In addition, these two antibodies are able to inhibit cell adhesion to recombinant and weakly modified OPN. The antibodies described herein may prove useful in determining the presence of modifications at specific sites and for identifying structural forms of OPN. Also, the sensitivity of these antibodies to PTMs suggests that caution must be taken when choosing anti‐OPN monoclonal antibodies to detect this highly modified protein. J. Cell. Biochem. 102: 925–935, 2007.

Increased Expression of the Remodeling- and Tumorigenic-Associated Factor Osteopontin in Pyramidal Neurons of the Alzheimer's Disease Brain

Osteopontin (OPN) is a glycophosphoprotein expressed by several cell types and has pro-adhesive, chemotactic, and cytokine-like properties. OPN is involved in a number of physiologic and pathologic events including angiogenesis, apoptosis, inflammation, oxidative stress, remyelination, wound healing, bone remodeling, cell migration and tumorigenesis. Since these functions of OPN, and the events that it regulates, are involved with neurodegeneration, we examined whether OPN was differentially expressed in the hippocampus of the Alzheimers disease (AD) compared with age-matched (59-93 years) control brain. We report for the first time the immunocytochemical localization of OPN in the cytoplasm of pyramidal neurons. In AD brains, there was a significant 41 % increase in the expression of neuron OPN compared with age-matched control brain. No staining of other neuronal cell types was observed. Additionally, there was a significant positive correlation between OPN staining intensity and both amyloid-beta load (r(2) = 0.25; P < 0.05; n = 20) and aging (r(2) = 0.32; P < 0.01; n = 20) among all control and AD subjects. Controlling for age indicated that OPN expression was significantly influenced by amyloid-beta load, but not age. While the functional consequences of this amyloid-beta associated increase in OPN expression are unclear, it is notable that OPN is primarily localized to those neurons that are known to be vulnerable to AD-related neurite loss, degeneration and death. Given that the induction of OPN expression (and amyloid-beta generation) is associated with remodeling and tumorigenesis, our results suggest that OPN may play a role in the aberrant re-entry of neurons into the cell cycle and/or neuronal remyelination in AD.

Osteopontin Traffic in Hypoxic Renal Epithelial Cells

Osteopontin (OPN), a secretory RGD-containing phosphoprotein, is induced in acute renal injury where it plays a renoprotective role. To investigate in depth the mode of OPN secretion under stress conditions, we analyzed OPN traffic in human renal proximal tubular epithelial cells (RPTEC). Western blot analysis and fluorescence microscopy revealed trace amounts of OPN in intact cells, whereas cytoplasmic OPN levels were significantly increased after 24–48 h hypoxia. Immunoelectron microscopy of RPTEC showed predominantly apical localization of gold-labeled OPN under normal conditions. Hypoxia (24 h) increased 2.5-fold immunodetectable gold-labeled OPN at the apical plasma membrane; further reoxygenation (2 h) augmented apical and basolateral labeling 2- and 10-fold, respectively. Analysis of apical and basolateral medium conditioned by RPTEC grown on semipermeable membranes using a specially developed ELISA showed a global decrease in secreted OPN after hypoxia, which recovered following 2 h reoxygenation. Agents known to disrupt the function of the Golgi apparatus (brefeldin A, monensin) or actin cytoskeleton (cytochalasin B) significantly inhibited OPN-GFP secretion in normoxic cells. In cells recovering from hypoxia, however, OPN secretion required functional Golgi apparatus, but was not affected by cytochalasin B. These findings demonstrate that stress inhibits OPN secretion by the process dependent on the functional Golgi apparatus and actin cytoskeleton; recovery restores OPN secretion, although its polarity may become perturbed.