A Review on the Role of Oxidative Stress and Inflammation in Necrotizing Enterocolitis and Benefits of the Phosphodiesterase Inhibitor Pentoxifylline

INTRODUCTION

Necrotizing enterocolitis: A highly activated inflammatory response resulting in disruption of intestinal epithelium and bowel necrosis is known as Necrotizing Enterocolitis (NEC). NEC is considered as one of prevalent emergency conditions mostly occurring in premature neonates. It affects approximately 10% of premature infants<1500 g (Claud, 2012), accompanied with high mortality and/or morbidity rate. Some of known common symptoms are diarrhea, feeding intolerance, abdominal distention and bloody stools. Alongside the aggravation of inflammation, intestinal perforation, peritonitis and systemic hypotension occur and patients require intensive medical care. Regardless of various attempts, the exact cause of NEC still remains challenging. However, some of the pathophysiological mechanisms that have been suggested so far are illustrated in the Fig. 2-4. However three main mechanisms can be described.

The first symptom of NEC is a reduction of blood flow in the intestine, which results in intestinal ischemia. This hypoxia condition is usually made by oxygen-derived free radicals and hydroxyl radicals by the mediation of xanthine oxidase.


Fig. 1:	Structure of pentoxifylline (C₁₃H¹⁸O₃)

These substances, besides leukocyte-derived free radicals, altogether, lead to an oxidant-mediated lipid peroxidation injury which is responsible for necrosis of intestine. Additionally, free radicals may increase vascular permeability and release of prostaglandins and leukotrienes (Okur et al., 1995; Erdener et al., 2004; Shah and Sinn, 2012; Caplan et al., 1990; Harpavat et al., 2012; Parks and Granger, 1983; Ciuffetti et al., 1991; Rezaie et al., 2007) (Fig. 2).

Another involved factor is over-activity of the immune system and an imbalance between activated pro-inflammatory responses with anti-inflammatory protection.


Fig. 2:	Intestinal ischemia as a pathophysiological mechanism in NEC, GI: Gastrointestinal; PG: Prostaglandins


Fig. 3:	Over-activity of immune system in NEC, NEC: Necrotizing enterocolitis; TLR: Toll like receptor; TNF: Tumor necrosis factor

Consequently, the augmented cytokine release, the increased level of tumor necrosis factor alpha (TNF-α), as a key factor for inflammation cascades and an activation of toll like receptor-4 (TLR-4) have been demonstrated to contribute in the aggravation of the inflammatory response in the intestine (Travadi et al., 2006; Arciero et al., 2013; Uguralp et al., 2004; Guven et al., 2009a) (Fig. 3).

Furthermore, reduced production of mucus besides the alteration of mucosal defenses and intestinal microbiota may bring about infection and bacterial translocation which contributes to gut injury via disruption of intestinal integrity.


Fig. 4:	The role of intestinal microbiota in development of NEC, GI: Gastrointestinal; NEC: Necrotizing enterocolitis

Inducible nitric oxide (iNOS) is also thought to induce enterocytes apoptosis that impairs mucosal barrier and bacterial translocation (Luedtke et al., 2012; Ergun et al., 2007) (Fig. 4).

Considering the relatively high mortality and morbidity of NCE, its prevention and treatment has become a field of interest in many clinical and experimental investigations (Patole, 2007). Although the exact pathophysiology of NEC is still unclear, due to the associated multifactorial causes and risk factors, various regimens have been successfully used. Modulation of inflammation by Platelet Activating Factor (PAF), interferon (INF) and glucocorticoids (Harpavat et al., 2012; Patole, 2007) are among the options. In addition, Ptx, vasoactive substances (Harpavat et al., 2012) and reducing the vascular permeability are among therapeutic strategies for the management of NEC (Parks and Granger, 1983). Targeting and modulation of the intestinal defenses, dietary and bacterial factors is effective in NEC treatment (Lin and Stoll, 2006). Several antibiotics have been used in addition to probiotics. Today, probiotics have been demonstrated to prevent development of NEC (Shah and Sinn, 2012; Patole, 2007).

To find more substances with beneficial effects, several investigations have been performed on experimental models. The details are summarized in the Table 1.

PENTOXIFYLLINE

Ptx (Fig. 1) is a tri-substituted purine (1-(5-oxohexyl)-3,7-dimethylxanthine) methyl xanthine derivative that besides its phosphodiesterase inhibitory effect, improves blood rheology and viscosity by inhibiting platelet aggregation and increasing blood cells flexibility. Phosphodiesterase inhibitors have been demonstrated to be effective in different inflammatory conditions and protect cells from oxidative-stress (Freitas and Filipe, 1995; Badri et al., 2011; Rahimi et al., 2010; Ghiasi et al., 2012).

In addition, Ptx exhibits immunomodulatory effects via decreasing the synthesis of TNF-α, interlukin-6 (IL-6), INF-γ and other pro-inflammatory cytokines throughout mucosal injury or sepsis. In the recent years, different phosphodiesterase inhibitors have been found beneficial in inflammatory bowel disease (Travadi et al., 2006; Patole, 2007; Harris et al., 2010; Lauterbach et al., 1999; Haque and Pammi, 2003; Salari and Abdollahi, 2012; Salari-Sharif and Abdollahi, 2010).

Ptx exerts its anti-oxidant properties by scavenging hydroxyl radicals and xanthine oxidase inhibition (Erdener et al., 2004; Salari and Abdollahi, 2012; Rezvanfar et al., 2012; Khoshakhlagh et al., 2007). It is believed to inhibit the generation of free radicals from leukocytes and the release of reactive oxygen metabolites during peripheral ischemia (Ciuffetti et al., 1991).

In this respect, Ptx has been used in several inflammatory infectious and vascular diseases (Harris et al., 2010; Rezvanfar et al., 2012). In addition, Ptx has been used in several clinical studies as a treatment option in shock and sepsis. Although, Haque and Pammi (2011) observed that Ptx may decrease the duration of disease in infants with sepsis but it does not affect the risk of development and incidence of NEC (Haque and Pammi, 2011). Some other reports demonstrate an increased incidence of NEC in placebo group of infants with sepsis which did not receive Ptx (Lauterbach et al., 1999; Lauterbach and Zembala, 1996).

Ptx has also been introduced as a therapeutic option to reduce the mortality and morbidity in premature infants with NEC (Harris et al., 2010). Given the immunomodulatory effects and demonstrated anti-oxidant and free radical scavenging effects of Ptx, it may comprise considerable potential to be in the package for management of NEC (Table 2).

METHODS

In this stduy we have reviewed the pathophysiology of NEC and evidences for the beneficial effect of anti-oxidant therapy and Ptx in this disease. In order to collect all evidences including experimental investigations and clinical trials, we used broad databases such as Google Scholar, PubMed, Scopus and Web of Science without date limitations.

RESULTS AND EVIDENCES

As summarized in Table 1, several studies have established animal models of ischemia and oxidative stress to induce NEC in the rat.

Table 1:	Experimental studies on the efficacy of several drugs on NEC

HB-EGF: Heparin-binding epidermal growth factor-like growth factor, HBO: Hyper baric oxygen, INF: Interferon, LBS: Lipopolysaccharide, MDA: Malondialdehyde, MPO: Myeloperoxidase,NAC: Acetylcysteine, NEC: Necrotizing enterocolitis, TAS: Total antioxidant status and XO: Xanthine oxidase

Table 2:	Details of experimental studies using Ptx for treatment of NEC

IR: Ischemia reperfusion, NEC: Necrotizing enterocolitis, Ptx: Pentoxifylline, TBARS: Lipid peroxidation, XO: Xanthine oxidase

These models mainly included different phases of hypoxy-reoxygenation, cold stress and/or lipopolysaccharide administration besides the enteral formula feeding. Consequently, the similar inflammatory condition in NEC occurs by lipid peroxidation injury and free radical release (Rezaie et al., 2007). In consideration of acquiring the main mediators and essential therapeutic targets, different interventions have been evaluated in NEC models. There are a substantial focus on substances with anti-oxidant activities such as N-acetylcysteine, vitamin E, omeprazole and hyperbaric oxygen (HBO. The results demonstrated that anti-oxidant therapy had protective effect on the intestinal injury and is able to diminish the severity of bowel damage in NEC. This beneficial effect was through inhibition of free radical formation (vitamin E, omeprazole) (Okur et al., 1995; Cadir et al., 2008), reduction of xanthine oxidase, malondialdehyde (MDA) levels and myeloperoxidase activity in addition to increase in total anti-oxidant status (N-acetylcysteine) (Ozdemir et al., 2012; Tayman et al., 2012) and improvement of white blood cell actions besides angiogenesis effects (HBO therapy) (Guven et al., 2009b). Some investigated treatments exhibited both anti-oxidant effect and anti-inflammatory protection like melatonin (Guven et al., 2011) and ozone therapy (Guven et al., 2009a). In addition, INF-alpha showed a modulator effect on cytokines resulting in reduction of the severity of NEC damage (Uguralp et al., 2004). Heparin-binding epidermal growth factor-like growth factor (HB-EGF) has been evaluated by Feng et al. (2006a; b) that showed preservation of gut barrier integrity and reduction of apoptosis. Furthermore, captopril reduced the mesenteric ischemia via inhibition of rennin-angiotensin system during shock (Zani et al., 2008).

DISCUSSION

NEC is known as one of the causes of death in neonate intensive care units that is prevalent in premature newborns. Prematurity, alteration in gastrointestinal microbial flora or infection, history of use of broad spectrum antibiotics, enteral feeding and sepsis are among proposed pathophysiologies (Luedtke et al., 2012; Hsueh et al., 2003). The maternal breast milk and conservative feeding practices are suggested as one of preventive factors to reduce NEC prevalence by influencing the colonization of microbial flora (Luedtke et al., 2012). Amongst investigated substances, Ptx may be of great interest based on results of experimental studies (Table 2) and due to its beneficial potentials for NEC management. Nevertheless, the safety profile of Ptx should be noted especially when used in combination with other drugs in the management of sepsis in neonates. Haque and Pammi (2003; 2011) showed that Ptx in combination with antibiotics reduces the mortality in affected infants with sepsis without leaving any adverse effects. Ptx does not result in any cardiac or bronchodilatory side effects when used in therapeutic doses (Harris et al., 2010). It caused no thrombocytopenia and bleeding in infants with sepsis or NEC (Harris et al., 2010). Additionally, Downard et al. (2012) investigated the direct peritoneal resuscitation method in an animal model of NEC. They suggested that this method could be effective in improving the blood flow in the bowel of affected animals. Thus, Ptx has the potential to preserve microvascular blood flow and can be considered as a preference in management of NEC. Collectively, further controlled clinical trials should be established to confirm the beneficial preventive and/or therapeutic use of Ptx in NEC, considering its safety issues in neonates as current evidences are mostly based on experimental reports and no clinical trial has been performed in NEC yet.

ACKNOWLEDGMENTS

This study is the outcome of an in-house financially non-supported study. Authors thank help of National Elite Foundation and Iran National Science Foundation (INSF).