**2.2 Peripheral vascular disease and ischemia**

Hypoperfusion (ischemia) resulting from peripheral vascular disease contributes to DFU development due to reduced oxygen supply to the diabetic foot [40]. The vascular disease in diabetic patients is mostly localized in arteries of the lower limb, with dismal prognosis [41, 42]. A major risk factor for the development of peripheral vascular disease is atherosclerosis [43, 44], a condition in which an artery become hardened or thickened due to a buildup of plaque in its inner lining. Atherosclerosis appears at a younger age in diabetic individuals and progresses more rapidly than in nondiabetic individuals [45, 46], thus making diabetes mellitus one of the risk factors for developing atherosclerosis. In the diabetic microcirculation, there is existence of structural changes, most notably, a thickening of the capillary basement membrane and endothelial dysfunction [47, 48]. As already mentioned, DPN reduces the production of neuropeptides which aid in wound healing [33–35]. These neuropeptides directly and indirectly (through mast cell release of histamine) cause increased permeability and vasodilation [49, 50]. The endothelium plays an important role in blood vessel wall function by synthesizing and releasing vasodilators such as prostaglandins and nitric oxide, which modulate vascular tone in pedal circulation [51, 52]. A host of mechanisms responsible for vascular dysfunction in diabetes have been identified to include over-production of reactive oxygen species, impaired nitric oxide pathway, abnormal production of vasoconstrictor prostanoids, intracellular signaling and advanced glycated end products [53–55]. This pathological vascular alteration in the diabetic foot contributes to DFU development, suggesting that diabetic patients must undergo a timely vascular examination for timely revascularization.

### **2.3 Infection**

The immune system of healthy people is much more robust and stronger than that of diabetic individuals [56, 57] and thus, foot infections in the latter group

need to be closely monitored and managed appropriately [58, 59]. Unsurprisingly, it is estimated that about 50% of patients with DFU present clinical signs of infection (either locally or systemically) [60, 61]. A foot infection is characterized by the presence of purulent secretions or at least two of the classic signs of inflammation (erythema, hyperemia, edema, or swelling and pain) [62, 63]. Patients with poor metabolic or uncontrolled diabetes are commonly afflicted with bacterial infection in which a lack of granulation tissue, delayed healing, appearance of necrotic tissue on wound surface and/or odor are prominent [64, 65]. Furthermore, these patients display a loss of sensitivity (i.e. diabetic peripheral neuropathy) in the affected foot due to damage caused to short and long fibers (A-beta and A-delta) secondary to hyperglycemia [66, 67]. Chronic hyperglycemia, a hallmark of diabetes mellitus, provides a suitable medium for bacterial growth, mainly aerobic Gram-positive cocci (e.g. *Staphylococcus aureus*), Gram-negative aerobes (e.g. *Pseudomonas aeruginosa*) and drug-resistant bacteria such as vancomycinresistant *S. aureus* and methicillin-resistant *S. aureus*, which were the common microorganisms identified in the diabetic foot [68–70]. In extreme situations, the bacterial infection of soft tissue spreads to the bones (causing osteomyelitis; an inflammation in the bone) and other surrounding tissues and organs. Unsuccessful attempts at halting the spread of the infection may lead to septicemia and amputation of the affected limb [65, 70]. Also, uncontrolled diabetes mellitus impairs the proliferation of fibroblasts and collagen synthesis while prolonging an inflammatory cascade [71, 72], a condition which compromises the wound healing abilities of diabetic patients. In summary, persistent hyperglycemia in diabetic individuals creates a suitable microenvironment for the growth of pathogenic bacteria, which contributes to DFU development and progression.
