Diabetic kidney disease (DKD) is the leading cause of CKD and ESKD in the United States and worldwide. Pharmacotherapy and lifestyle modifications for glycemia, dyslipidemia, and BP control have shown success in slowing the progression of DKD. Traditional treatments, such as angiotensin-converting enzyme inhibitors or angiotensin receptor blockers and more recently the use of sodium-glucose cotransporter 2 inhibitors, nonsteroidal selective mineralocorticoid receptor antagonists, such as finerenone, and glucagon-like peptide 1 receptor agonists, have led to added benefits on various outcomes. However, significant residual risk for DKD progression remains despite the current standard-of-care approaches. Arteriolar hyalinosis (AH) is among the key findings seen on kidney biopsies of patients with DKD. It results from the excessive accumulation of hyaluronan (HA) in the arterioles. AH has not been targeted specifically by any of the therapeutic methods currently being used. We discuss in this manuscript the potential use of a selective therapy targeting AH and the increased total renal HA deposits using a HA synthesis inhibitor in DKD.

The role of hyaluronan (HA) in the development and progression of diabetic kidney disease (DKD), as well as the precise mechanisms and consequences of HA involvement in this pathology are still to be clarified.

In this study, we assayed the effects of the HA synthesis inhibitor 4-methylumbelliferone (4-MU) on the development of DKD. Diabetic type 2 model mice (eNOS^-/- C57BLKS/J^db) were fed artificial diets containing 5% 4-MU or not for 9 weeks. Plasma glucose, glomerular filtration rate (GFR), albumin to creatinine ratio (ACR), and biomarkers of kidney function and systemic inflammation were measured at baseline and after treatment. Diabetic nephropathy was further characterized in treated and control mice by histopathology.

Treated animals consumed a daily dose of approximately 6.2 g of 4-MU per kg of body weight. At the end of the experimental period, the 4-MU supplemented diet resulted in a significant decrease in non-fasting plasma glucose (516 [interquartile range 378-1170] vs. 1149 [875.8-1287] mg/dL, P=0.050) and a trend toward lower HA kidney content (5.6 ± 1.5 vs. 8.8 ± 3.1 ng/mg of kidney weight, P=0.070) compared to the control diet, respectively. Diabetic animals treated with 4-MU showed significantly higher GFR and lower urine ACR and plasma cystatin C levels than diabetic controls. Independent histological assessment of DKD also demonstrated a significant decrease in mesangial expansion score and glomerular injury index in 4-MU-treated mice compared to controls. Plasma glucose showed a strong correlation with kidney HA levels (r=0.66, P=0.0098). Both total hyaluronan (r=0.76, P=0.0071) and low-molecular-weight hyaluronan content (r=0.64, P=0.036) in the kidneys correlated with urine ACR in mice.

These results show that the hyaluronan synthesis inhibitor 4-MU effectively slowed the progression of DKD and constitutes a potential new therapeutic approach to treat DKD.

Accumulation of extracellular matrix (ECM) components is an early sign of diabetic nephropathy. Also the glycosaminoglycan hyaluronan (HA) is elevated in the renal interstitium during experimental diabetes. The mammalian target of rapamycin (mTOR) pathway participates in the signaling of hyperglycemia-induced ECM accumulation in the kidney, but this has not yet been investigated for HA. We hypothesized that interstitial HA accumulation during diabetes may involve mTOR activation.

Diabetic rats (6 weeks post-streptozotocin (STZ)) were treated with rapamycin to inhibit mTOR or vehicle for 2 additional weeks. Kidney function (glomerular filtration rate, renal blood flow, urine output) and regional renal HA content were thereafter analyzed. The ability of the animals to respond to desmopressin was also tested.

Diabetic animals displayed hyperglycemia, proteinuria, hyperfiltration, renal hypertrophy, increased diuresis with reduced urine osmolality, and reduced weight gain. Cortical and outer medullary HA was elevated in diabetic rats. Urine hyaluronidase activity was almost doubled in diabetic rats compared with controls. The ability to respond to desmopressin was absent in diabetic rats. Renal blood flow and arterial blood pressure were unaffected by the diabetic state. In diabetic rats treated with rapamycin the proteinuria was reduced by 32%, while all other parameters were unaffected.

Regional renal accumulation of the ECM component HA is not sensitive to mTOR inhibition by rapamycin, while proteinuria is reduced in established STZ-induced diabetes. Whether the diabetes-induced renal accumulation of HA occurs through different pathways than other ECM components, or is irreversible after being established, remains to be shown.

The glycosaminoglycan (GAG) hyaluronan (HA) is recognized as an important structural component of the extracellular matrix, but it also interacts with cells during embryonic development, wound healing, inflammation, and cancer; i.e., important features in normal and pathological conditions. The specific physicochemical properties of HA enable a unique hydration capacity, and in the last decade it was revealed that in the interstitium of the renal medulla, where the HA content is very high, it changes rapidly depending on the body hydration status while the HA content of the cortex remains unchanged at very low amounts. The kidney, which regulates fluid balance, uses HA dynamically for the regulation of whole body fluid homeostasis. Renomedullary HA elevation occurs in response to hydration and during dehydration the opposite occurs. The HA-induced alterations in the physicochemical characteristics of the interstitial space affects fluid flux; i.e., reabsorption. Antidiuretic hormone, nitric oxide, angiotensin II, and prostaglandins are classical hormones/compounds involved in renal fluid handling and are important regulators of HA turnover during variations in hydration status. One major producer of HA in the kidney is the renomedullary interstitial cell, which displays receptors and/or synthesis enzymes for the hormones mentioned above. During several kidney disease states, such as ischemia-reperfusion injury, tubulointerstitial inflammation, renal transplant rejection, diabetes, and kidney stone formation, HA is upregulated, which contributes to an abnormal phenotype. In these situations, cytokines and other growth factors are important stimulators. The immunosuppressant agent cyclosporine A is nephrotoxic and induces HA accumulation, which could be involved in graft rejection and edema formation. The use of hyaluronidase to reduce pathologically overexpressed levels of tissue HA is a potential therapeutic tool since diuretics are less efficient in removing water bound to HA in the interstitium. Although the majority of data describing the role of HA originate from animal and cell studies, the available data from humans demonstrate that an upregulation of HA also occurs in diabetic kidneys, in transplant-rejected kidneys, and during acute tubular necrosis. This review summarizes the current knowledge regarding interstitial HA in the role of regulating kidney function during normal and pathological conditions. It encompasses mechanistic insights into the background of the heterogeneous intrarenal distribution of HA; i.e., late nephrogenesis, its regulation during variations in hydration status, and its involvement during several pathological conditions. Changes in hyaluronan synthases, hyaluronidases, and binding receptor expression are discussed in parallel.

Sulfated glycosaminoglycan synthesis by human diabetic, non-diabetic, and embryonic cells was studied. No effect of insulin on net synthesis was noted. Thus, the data do not indicate a role for total sulfated glycosaminoglycan production in the diabetic connective tissue disturbance mediated by insulin.

Unilateral renal ischemia during 30 min causes severe, non-reversible renal damage in diabetic (DM) rats, but not in nondiabetic rats. Hyaluronan (HA) is a glycosaminglycan involved in various forms of renal injury. We examined the role of HA and CD44, a major receptor for HA, in the development of postischemic renal injury in DM rats.

The left renal artery of streptozotocin diabetic Wistar rats was clamped for 30 min. The HA content in the kidneys was measured. A biotinylated HA-binding probe was used to localize HA. Inflammatory cells and other cells expressing CD44 were identified by immunohistochemistry.

In ischemic DM kidneys the renal HA-content started to increase already after 24 h and significantly so after 1-8 weeks after ischemia/reperfusion (I/R). The relative water content of the kidneys increased in parallel. HA started to appear in the cortex of ischemic DM kidneys 1 week after I/R. In contrast, the non-DM ischemic kidneys showed no increase of HA and water content after 1-8 weeks after I/R. The tubular cells in the cortex and outer medulla demonstrated increased staining for CD44. In the same compartments the increased numbers of infiltrating inflammatory cells also expressed CD44.

HA-accumulation in the renal cortex might contribute to the renal damage seen after transient ischemia in DM rats by promoting inflammation through interaction between HA and CD44 expressing inflammatory cells. Furthermore, HA accumulation may contribute to an interstitial renal edema.

GAG metabolism was investigated in rats with experimentally induced diabetes. In comparison to control animals, the uptake of 35S-sulfate was diminished in tissues of diabetic animals. Streptozotocin-induced diabetes showed a significant decrease in the content of GAG fractions except that of non-sulfated GAG in liver and kidney which was unchanged as compared to the control group. In rats rendered diabetic by alloxan, non-sulfated GAG increased appreciably in liver and kidney whereas highly sulfated GAG remained unchanged. In the skins of alloxan-diabetic rats both total and sulfated GAG decreased significantly. The activities of liver beta-glucuronidase, beta-N-acetyl glucosaminidase and cathepsin D were significantly increased in rats treated with streptozotocin and alloxan. In streptozotocin-diabetic rats, renal beta-glucuronidase and beta-N-acetyl glucosaminidase activities were reduced while cathepsin D activity was similar to that of controls. The renal beta-N-acetyl glucosaminidase and cathepsin D activities of alloxan-treated rats were not significantly different from normal but their beta-glucuronidase was significantly increased. In the spleen of streptozotocin-diabetic rats all the enzymes were increased except beta-N-acetyl glucosaminidase which remained unaltered. Increased excretion of uronic acid was observed in diabetic groups. These results collectively indicate that both streptozotocin- and alloxan-induced diabetes altered the synthesis and catabolism of GAG.

One of the important substances in the ground substance is acidic glycosaminoglycans (AGAGs). Changes of AGAGs in the inner ear and in other organs were investigated using alloxan diabetic mice in order to contribute to the understanding of diabetic hearing impairment. In the diabetic group, gradual increases of AGAGs were observed in each tissue. On the 60th day after alloxan injection, AGAG values were increased 3.5-fold in the cochlea, 2.5-fold in the brain, 13-fold in the liver, twofold in the kidney, and fivefold in the pancreas compared with the control values. It is interesting to note that both the cochlea and pancreas showed continuous increases of AGAGs.

To assess the effects of streptozotocin-induced diabetes on the substrates utilized in the formation of glycoproteins, the pools of uridine 5'-diphosphoglucose (UDPG), uridine 5'-diphosphogalactose (UDP-GAL), uridine 5'-diphosphoglucuronic acid (UDPGA), and uridine 5'-diphospho N-acetyl galactosamine (UDPA-GAL) were measured in the renal cortex of control and over a 48-hr period in diabetic rats. In control rats these pools measured: UDPG, 256 +/- 23; UDP-GAL, 75 +/- 14; UDPGA, 147 +/- 16; UDPAG, 367 +/- 23; UDPG-GAL, 131 +/- 13 nmoles/mg DNA. In diabetic rats, except for UDP-GAL, all pools were increased 41 to 68%. The incorporation of radiolabeled orotate was increased in all pools, except UDP-GAL, in diabetic rats by 41 to 77% compared to control rats. The incorporation into UDPG and UDPAG was increased even after correction for the specific radioactivity of their immediate precursor, uridine 5'-triphosphate (UTP). Expansion of the UTP pool after orotate infusion was associated with an increase in the size of the UDPG and UDPAG pools in both control and diabetic rats. Depletion of the UTP pool after adenine infusion in controls was associated with a decrease in all pools. This study demonstrates that after the induction of diabetes there is a rapid increase in the bioavailability of substrates utilized in the synthesis of glycoproteins and glycosaminoglycans. It is theorized that this increase is necessary for the augmented synthesis of basement membrane-like material in the diabetic kidney.

Eighteen diabetic children aged between 8.5 and 16.5 years (mean 12.5 years) who had been diabetic for 1 to 10 years (mean 4.1 years) were examined for their urinary glomerular basement membrane (GBM) antigen excretion by means of immunoelectrophoresis and for alterations of the retinal vessels by fluorescence angiography. None of these patients showed albuminuria or hypertension. As compared to 40 healthy controls aged between 5 and 17 years, altered GBM antigen mobility (alpha-1) was found in 9 out of these 18 diabetics, whereas the remaining 9 children had normal GBM antigen mobility (alpha-2). Pathological fluorescence angiography findings on the other hand were evident in 7 children with altered GBM mobility, but only in 4 diabetics with normal GBM antigen mobility. This trend reflects the similarity of biochemical and functional characteristics of basement membranes in the retinal and kidney vessels supporting the well established association of vascular changes in both organs in patients with diabetes mellitus. GBM antigen excretion into urine could be useful for detecting early microvascular alterations in the kidneys in juvenile diabetics where diagnosis of early glomerulosclerosis is important.

Glomerular basement membrane antigens excreted into urine, urinary acid glycosaminoglycans and hemoglobin AIc plasma levels were estimated in 18 diabetic children. In comparison to controls we detected significant differences, in that diabetics with changed (alpha-1-) mobility on immunoelectrophoresis revealed increased urinary acid glycosaminoglycan excretion (p less than 0.01) and elevated HbAIc concentrations in plasma (p less than 0.01). HbAIc levels and urinary glycosaminoglycans correlated significantly (r = 0.7, p less than 0.001). Increased glycosaminoglycans as well as increased blood glucose reflected by elevated HbAIc could lead to the biochemical glomerular basement membrane changes found on immunoelectrophoresis. The results suggest a metabolic hypothesis for the development of diabetic glomerulopathy.

Acidic glycosaminoglycans in the ground substance of the membranous cochlear lateral wall, kidney, brain and liver of the guinea pigs were quantitatively and qualitatively analyzed, using electrophoretic microanalysis. The lateral wall of the cochlea showed highest content (0.46 +/- 0.08%/D.W.) of acidic glycosmainoglycans, which were chondroitin sulfate-B (75%), chondroitin sulfate-A (12%) and hyaluronic acid (13%). However, the pattern of these acidic glycosaminoglycans in the inner ear differed from the other organs. A possible role of acidic glycosaminoglycans in the inner ear was discussed.

The acidic glycosaminoglycans (AGAG) in normal human kidneys were fractionated on Dowex 1-X2 columns and analysed by electrophoretic separation in three buffers on cellulose acetate membranes and gel filtration on Sephadex G-100 columns, before and after digestion with chondroitinases and streptomyces hyaluronidase. Thin-layer chromatography was also performed to separate glucosamine from galactosamine moieties. Enzymatic digestion combined with electrophoretic characterization indicated that heparan sulfates exist as the main AGAG which accounted for two-fifths of the total AGAG. Hyaluronic acid and dermatan sulfates accounted for one-fourth and one-sixth of the total kidney AGAG, respectively. Chondroitin sulfate isomers (4-sulfate and 6-sulfate) consisted of the residual one-sixth of the total AGAG. An oversulfated chondroitin sulfate was detected in a small amount by demonstration of the unsaturated disulfated disaccharide after digestion with chondroitinase-ABC but not with chondroitinase-AC.

Connective tissue macromolecules, glycosaminoglycans, glycoproteins, collagen, and elastin were isolated from different parts of the respiratory system and characterized. The materials included bronchiolar tissue, gas-exchange tissue, lung pleura, and tracheal mucosa. The similarity of the macromolecular composition of lung pleura and tracheal mucosa suggests a common cellular component in these structures. The high concentration of GAG and collagen in bronchiolar tissue is consistent with the cartilagenous nature of this tissue. Particularly interesting is the high content of heparin in all pulmonary structures, a relatively greater content of hyaluronic acid in gas-exchange tissue, and a high content of heparan sulfate and dermatan sulfate in vascular tissues. Elastin also occurs as a major fibrous structure. Although the biologic role of these connective tissue macromolecules has not been established, certain functional relationships are inferred.