**9. Conclusions**

A growing body of evidence, in in vitro and in vivo model systems, implicates intracellularly generated oxidative stress as one of the contributing factors in the cascade of events leading to cytokine-induced metabolic dysfunction and demise of the islet beta cell. In this review, I attempted to summarize the known evidence in support of regulatory roles of Noxs, specifically Nox1 and Nox2, in cytokine-induced alterations in the metabolic functions of the islet. Evidence in favor of a regulatory role for Nox4 is emerging, but potential roles of Nox3 and Nox5 in this signaling cascade remain relatively poorly understood. Several pharmacological approaches have been employed to decipher the roles of Noxs, iNOS, and 12-LO as mediators of cytokine-induced metabolic dysregulation of the islet beta cell (Table 1). Indeed, such approaches have provided useful insights into these pathways as potential targets to prevent/halt the metabolic defects. Potential regulatory roles of PN as a mediator of beta cell damage under the duress of cytokines remains to be examined further. In addition, much is unknown with regard to signaling mechanisms and regulatory proteins/factors that promote crosstalk between iNOS-12-LO-Nox2 signaling pathways in the cascade of events leading to cytokine dysregulation of the islet beta cell. These aspects need to be addressed further.


**Table 1.** Inhibitors of the Nox, iNOS, and 12-LO signaling pathways employed in studies highlighted in this review.

Based on the evidence, principally accrued in our laboratory on the regulatory roles of small G proteins in the generation of iNOS-derived ROS and Nox2-derived ROS, I propose a model for a potential crosstalk between these signaling pathways in eliciting damage to the rat islet beta cell following exposure to proinflammatory cytokines (Figure 3). Briefly, using pharmacological approaches, it was demonstrated that IL-1β-induced iNOS expression and NO release are under the control of H-Ras, a small G protein belonging to the Ras superfamily of G proteins [80,81]. These findings were further confirmed with bacterial toxins that promote glucosylation and inactivation of small G proteins [80,81]. Interestingly, data accrued from the investigations involving bacterial toxins suggested that activation of Rho G proteins (e.g., Rac1) is not a requisite for IL-1β-induced iNOS expression and NO acid release [81]. Subsequent studies involving specific inhibitors of Rac1 (e.g., NSC23766) further affirmed the postulation that Rac1 is involved in ROS generation but not NO production in a cytokine-challenged beta cell [53]. The combined effects of intracellularly generated NO (due to activation of iNOS) and ROS (due activation of Nox) could contribute to maximal damage of the mitochondrial membrane properties leading to metabolic dysfunction of the beta cell (Figures 2 and 3). It is emphasized that this model, involving H-Ras, may not be applicable to the human beta cells since they do not express the iNOS-NO signaling pathway, and ye<sup>t</sup> exert sensitivity to NO, which is secreted from activated immune cells. The overall concept of intracellular generation of PN and its effects on mitochondrial dysregulation, loss in GSIS, and apoptotic demise of the islet beta cell under the duress of cytokines was proposed earlier [85]. Additional studies are needed to further substantiate this formulation. It is my hope that the future advances in the field of Nox biology will help not only in our current understanding of this class of enzymes but also in the development of small-molecule compounds (via combinatorial chemistry approaches) with a high degree of specificity to inhibit the iNOS-Nox-LO pathways. The development of methodologies to boost the overall antioxidant capacity of the islet beta cell is also warranted. Lastly, novel approaches to accelerate detoxification of intracellularly generated H2O2 in specific subcellular compartments (e.g., cytosol and mitochondria) might prove valuable in preventing cytokine-mediated dysfunction and demise of the pancreatic islet beta cell.

**Figure 3.** Potential involvement of small G proteins in cytokine-induced NO release and ROS formation leading to mitochondrial dysfunction and demise of the rat islet beta cell.

**Funding:** This research is supported by a Merit Review (I01 BX004663) and a Senior Research Career Scientist (IK6 BX005383) awards from the US Department of Veterans Affairs, and a gran<sup>t</sup> from the National Eye Institute (EY022230). The author also received support from the Juvenile Diabetes Foundation International.

**Acknowledgments:** The author thanks the former and current members of his research group and collaborators for their contribution to the work highlighted herein.

**Conflicts of Interest:** The author declares no conflict of interest.
