Normalization of Intracellular Calcium: A Sweet Solution to Neutrophil Dysfunction in Diabetes?

The diapedesis of the white corpuscles, their migration through the vessel wall into the tissues, is one of the principal means of defense possessed by the animal.As soon as the infective agents have penetrated into the body, a whole army of white corpuscles proceed towards the menaced spot, there entering into a struggle with the microorganisms. The leukocytes, having arrived at the spot where the intruders are found, seize them after the manner of Amoebae and within their bodies subject them to intracellular digestion. Ilya I'litch Metchnikoff (1882) The increased susceptibility of persons with diabetes mellitus to complicated soft-tissue, urinary tract, and lower respiratory tract infections is likely to be at least partially due to deficient polymorphonuclear leukocyte (PMNL) function [1-3]. Independent of acidosis and uremia, PMNLs from patients with diabetes mellitus show impaired adherence [4-6], chemotaxis [7], phagocytosis [8, 9], oxidative activity [10], and bactericidal activity [8-10]. These defects are detectable with fasting blood glucose levels greater than 7.2 mmol/L and within 36 hours of insulin withdrawal [10]. Clinical studies have shown that improved PMNL function accompanies improvement of fasting blood glucose values [3], whereas in vitro studies indicate that such improvement occurs within 15 to 20 minutes of co-incubation with insulin [10]. The PMNL bactericidal defect in patients with diabetes against Staphylococcus aureus [9, 10] may be especially worthy of attention because S. aureus is a frequent and serious pathogen in these patients. [11], and PMNL defects are associated with an increased frequency of serious S. aureus infections [12]. In this issue, Alexiewicz and colleagues [13] report their study of the physiologic significance of increased cytosolic calcium ([Ca2+]i) concentrations in PMNLs from patients with non-insulin dependent diabetes before and after treatment with glyburide. Their results provide important insight about the mechanisms that may underlie PMNL dysfunction in patients with diabetes. Pretreatment PMNLs from these patients showed decreased phagocytic activity and increased resting [Ca2+]i levels when compared with PMNLs from controls; adenosine triphosphate (ATP) concentrations were also decreased in the PMNLs of patients with diabetes. After 3 months of glyburide therapy, serum glucose levels approached normal and hemoglobin A1c levels were significantly improved in these patients. These improvements were accompanied by significant normalization of [Ca2+]i levels, intracellular ATP concentrations, and an in vitro assay of the phagocytic activity in the PMNLs from these patients. A strong correlation among the degree of hyperglycemia, [Ca2+]i levels, and dysfunction of the phagocytic activity in PMNLs was noted. These results expand on previous studies by this group that showed that increases in [Ca2+]i levels in the PMNLs of patients with chronic renal failure are associated with decreased phagocytic activity [14] and that improvement of phagocytic activity is related to the normalization of [Ca2+]i levels rather than to normalization of intracellular ATP concentrations [15]. These findings raise the possibility that interventions capable of causing [Ca2+]i levels to return to normal, such as the administration of calcium channel blockers [16], may improve the bactericidal activity of PMNLs and thus decrease the rate of infection in persons with diabetes mellitus. Further development of such hypotheses depends on whether normalization of [Ca2+]i correlates with improvement in the other PMNL functions that are defective in patients with uncontrolled diabetes mellitus. Much as Metchnikoff described, effective PMNL antimicrobial activity depends on the successful integration of several cellular functions. Polymorphonuclear leukocytes must first adhere to capillary endothelial cells adjacent to the site of infection, then traverse the endothelium to reach the site of microbial proliferation. Only after polymorphonuclear leukocytes attach to the infecting microorganism can ingestion (phagocytosis) occur. Finally, microbial killing occurs after various toxic proteins and oxygen radicals are delivered to the phagosome [12]. Although chemotaxis, phagocytosis, and oxidative activity have all been found to be abnormal in PMNLs from persons with poorly controlled diabetes, Alexiewicz and colleagues [13] have thus far shown only that an improvement in phagocytic activity accompanies normalization of blood glucose and [Ca2+]i levels. Furthermore, the phagocytosis assay used by Alexiewicz and colleagues measures the association of complement- and endotoxin-coated oil red O particles with PMNLs, but it does not assess the actual cellular ingestion or IgG-dependent cellular activation [17]. Optimal ingestion and cellular oxidative activation depend on the ligation of both complement and IgG receptors [12]. Importantly, bacteria that adhere to PMNLs but are not ingested are largely unaffected by PMNL-mediated killing [18]. Thus, the clinical implications of the improvement of phagocytic activity as measured by these authors require further study. How might increased [Ca2+]i levels in PMNLs impair PMNL function? Normally, PMNL activation is dependent on a rapid, phospholipase C-mediated increase in [Ca2+]i levels from less than 100 nmol/L to micromolar concentrations [12]. Alexiewicz and colleagues have previously shown that, when compared with PMNLs from controls, the [Ca2+]i levels are elevated in resting PMNLs from patients who are receiving long-term hemodialysis and that these cells evince a smaller increase in [Ca2+]i levels after cellular stimulation [14]. Thus, an intriguing hypothesis is that the increased [Ca2+]i levels in resting PMNLs from patients with diabetes is also associated with a lesser increase of [Ca2+]i levels after cellular stimulation and that normalization of the resting [Ca2+]i is accompanied by normalization of [Ca2+]i kinetics. In yet another study, nifedipine partially corrected [Ca2+]i levels and ATP concentrations in PMNLs from patients with and without diabetes who were receiving hemodialysis and substantially improved the cellular phagocytic capacity as judged by the oil red O assay [18]. However, because increased [Ca2+]i levels in patients who receive hemodialysis is largely attributable to abnormalities in parathyroid hormone metabolism, extrapolation of these findings to PMNLs from patients with diabetes may not be appropriate. Furthermore, caution is in order because verapamil not only inhibits PMNL Ca2+ influx but also inhibits activation of PMNL oxidative metabolism [19]. There are many reasons for recommending tight control of glycemia in patients with diabetes. Intensive treatment of hyperglycemia reduces the incidence and severity of retinopathy, neuropathy, and nephropathy in patients with insulin-dependent diabetes mellitus [20]. Alexiewicz and colleagues [13] have now shown that PMNL dysfunction in patients with hyperglycemic, non-insulin dependent diabetes mellitus is associated with increased [Ca2+]i levels and that glycemic control is associated with both normalization of [Ca2+]i and improved PMNL phagocytic capacity. These findings illuminate the cellular biochemistry that may link hyperglycemia to decreased PMNL function, and they confirm previous clinical and laboratory observations that better glycemic control improves neutrophil function [5, 10]. Finally, it may be tantalizing to speculate that pharmacologic modulation of [Ca2+]i levels may improve PMNL function and reduce the risk for infection even in patients with persistent hyperglycemia. However, before such clinical interventions can be entertained, it remains to be shown that normalization of [Ca2+]i itself improves overall PMNL bactericidal activity.