Background

We recently reported on the enhanced tubular expression of two discrete isoforms of the MMP-2 (full length and N-terminal truncated, FL-MMP-2, NTT-MMP-2) in a murine model and human diabetic kidneys. In the present study, we examined in more detail the temporal and spatial distributions of MMP-2 isoform expression in murine models of Type 1 and Type 2 diabetes mellitus.

Methods

Diabetic models were streptozotocin (STZ)-induced diabetes (Type 1 diabetes mellitus) and db/db mice (Type 2 diabetes mellitus). We quantified the abundance of two isoforms of MMP-2 transcripts by qPCR. A spatial distribution of two isoforms of MMP-2 was analyzed semi-quantitatively according to time after injection of STZ and with increasing age of db/db mice. Furthermore, immunohistochemistry for nitrotyrosine was performed to examine a potential association between oxidative stress and MMP-2 isoform expression.

Results

Both isoforms of MMP-2 were upregulated in whole kidneys from STZ and db/db mice. In the case of FL-MMP-2, mRNA levels significantly increased at 12 and 24 weeks in STZ mice, while the isoform expression was significantly increased only at 16 weeks, in the db/db mice. FL-MMP-2 protein levels increased in the cortices and outer medullae of both STZ and db/db mice as a function of the duration of diabetes. For NTT-MMP-2, mRNA levels increased earlier at 4 weeks in STZ mice and at 10 weeks of age in db/db mice. The expression of NTT-MMP-2 also increased, primarily in the cortices of STZ and db/db mice, as a function of the duration of diabetes. Quantitatively, these findings were consistent with the qPCR results in the case of NTT-MMP-2, respectively (STZ 24 weeks, 3.24 ± 3.70 fold; 16 weeks db/db, 4.49 ± 0.55 fold). In addition, nitrotyrosine was expressed primarily in cortex as compared to medulla as a function of the duration of diabetes similar to NTT-MMP-2 expression.

Conclusions

Two isoforms of MMP-2 are highly inducible in two diabetic murine models and become more abundant as a function of time. As the expression patterns were not the same in the two isoforms of MMP-2, it is possible that each isoform has a discrete role in the development of diabetic renal injury.

Background

Diabetic cardiomyopathy (DM CMP) is defined as cardiomyocyte damage and ventricular dysfunction directly associated with diabetes independent of concomitant coronary artery disease or hypertension. Matrix metalloproteinases (MMPs), especially MMP-2, have been reported to underlie the pathogenesis of DM CMP by increasing extracellular collagen content.

Purpose

We hypothesized that two discrete MMP-2 isoforms (full length MMP-2, FL-MMP-2; N-terminal truncated MMP-2, NTT-MMP-2) are induced by high glucose stimulation in vitro and in an experimental diabetic heart model.

Methods

Rat cardiomyoblasts (H9C2 cells) were examined to determine whether high glucose can induce the expression of the two isoforms of MMP-2. For the in vivo study, we used the streptozotocin-induced DM mouse heart model and age-matched controls. The changes of each MMP-2 isoform expression in the diabetic mice hearts were determined using quantitative real-time polymerase chain reaction (qRT-PCR). Immunohistochemical stains were conducted to identify the location and patterns of MMP-2 isoform expression. Echocardiography was performed to compare and analyze the changes in cardiac function induced by diabetes.

Results

Quantitative RT-PCR and immunofluorescence staining showed that the two MMP-2 isoforms were strongly induced by high glucose stimulation in H9C2 cells. Although no definite histologic features of diabetic cardiomyopathy were observed in diabetic mice hearts, left ventricular systolic dysfunction was determined by echocardiography. Quantitative RT-PCR and IHC staining showed this abnormal cardiac function was accompanied with the increases in the mRNA levels of the two isoforms of MMP-2 and related to intracellular localization.

Conclusion

Two isoforms of MMP-2 were induced by high glucose stimulation in vitro and in a Type 1 DM mouse heart model. Further study is required to examine the role of these isoforms in DM CMP.