The systemic inflammatory response appears in the early phase of severe acute pancreatitis (SAP), which is determined by pancreatic and peripancre- atic necrosis. Exacerbation of the systemic inflammatory response results in immune function paralysis and life- threatening complications, even multiple organ failure. Therefore, attempts to reduce the excessive inflammatory response in SAP, for example, by blocking certain proin- flammatory cytokines or secondary mediators involved in these reactions, are important.1-4 However, there is no good strategy to attenuate the inflammatory response to date. Thus, it is necessary to find a new pathway to decrease hyperinflammation in SAP patients.

It is widely accepted that long-chain polyunsaturated fatty acids (PUFAs) not only provide energy but also possess important additional functions. While ω-6 fatty acids (FAs) have been demonstrated to enhance the inflammatory response, ω-3 FAs have been observed to suppress inflammation and ameliorate the course of infection, such as sepsis, ulcerative colitis, and acute respiratory distress syndrome.4-12 In general, eicosanoids derived from ω-3 FAs cause biologic responses opposite to those of ω-6 FAs, including less inflammatory response after trauma or surgery. These data indicate that these fatty acids have potential clinical benefit for patients undergoing intensive care.

Meanwhile, patients often need parenteral nutrition (PN) in the early phase of SAP because of increased metabolic demand or the presence of ileus. However, it is unclear whether addition of ω-3 FAs to PN influences inflammatory response in SAP patients. Thus, we designed a randomized, double-blind, and controlled study to compare the effects of ω-3 and ω-6 FAs emulsions in PN on the hyperinflammatory response and systemic disease sequelae in SAP patients. In the present study, we used soybean oil (SO) and fish oil (FO) emulsions as major sources of ω-6 and ω-3 FAs, respectively.

Methods

Patients

Forty patients suffering from severe acute pancreatitis were enrolled between April 2006 and February 2007. The inclusion criteria were based on the Summary of the International Symposium on Acute Pancreatitis, Atlanta, 1992. Acute pancreatitis is an acute inflammatory process of the pancreas, with variable involvement of other regional tissues or remote organ systems. Moreover, severe acute pancreatitis is associated with organ failure and/or local complications, such as necrosis, abscess, or pseudocyst.13 Patient characteristics are shown in Table 1. The study was approved by the Ethics Committee of the medical faculty of Jinling Hospital, and the followed procedures were in accordance with the Helsinki Declaration of 1975, as revised in 1989. Voluntary informed consent of each patient was obtained before the commencement of the investigation. Patients were enrolled within 72 hours after onset of SAP and were randomized to receive PN supplemented with either SO or SO-FO emulsion in a doubleblind manner (Table 2). Patients were 18-80 years old before entry to the study. The patients were observed for 5 days in the 1 7-bed intensive care unit (ICU) of the Department of General Surgery.6,7

Exclusion Criteria

Patients were excluded if admitted to the hospital after 72 hours of SAP onset. Patients who were pregnant, underweight or obese (body mass index <16 or >30 kg/m^sup 2^), or those with known alcohol or drug abuse were excluded from participation. Patients with hypertriglyceridemia, hyperthyroidism, chronic liver disease, HIV infection, hepatitis, severe cardiac or renal disease, or the use of insulin, corticoids, cytostatics, or cyclooxygenase inhibitors within 2 weeks prior to the study were also excluded.

Double Blinding and Randomization

Patients were assigned to the respective group by computerderived block randomization. The nurse was the only person aware of the randomization list. Accordingly, she prepared solutions in the Nutrition Center of Jinling Hospital for each patient. PN was then delivered blindly (with patient identification) to the ICU and further handled by a nurse who was unaware of the study protocol. Thus, the investigators were blinded to the infused drugs.

Nutrition Interventions

It was reported that ω-3 FAs- supplemented fat emulsion in PN for 5 days after surgery was clinically safe and led to alteration of the FA profile in plasma and attenuation of inflammation. Patients were randomly assigned to the 2 groups to receive isonitrogenous (0.2 g N/kg/d) and isoenergetic (117 kJ/28 kcal/kg/d) PN. The PN formula is shown in Table 2. It was administered by an indwelling central venous catheter. The regimen consisted of 1.25 g of amino acids/kg/d, 3 g of glucose/kg/d, and 1 g of fat/kg/d. Lipid was either SO (Lipovenos 20%; Fresenius, Germany) or FO-supplemented SO (Omegaven 10%; Fresenius, Germany). In the SO-FO group, the ω-6 FAs content of PN was partially replaced by ω-3 FAs up to 10 g daily, about 0.15 - 0.2 g/kg body weight, which was the recommended daily dosage. Thus, the ω-3/ω-6 FAs ratio was about 1:4 in the SO-FO group. A previous report found that this dosage was associated with attenuated inflammatory response.12 Moreover, the daily regimen contained fat-soluble (Vitalipid; Fresenius, Germany) and water-soluble (Soluvit; Fresenius, Germany) vitamins as well as trace elements (Addel N; Fresenius, Germany). No enteral or oral food intake was permitted in the 5 days.

Samples

On day 1 of PN, baseline values were obtained before PN was started (8:00 am), including blood samples. Then, blood samples were collected on day 6 after the completion of PN. For laboratory measurements, 15 mL of whole blood was withdrawn from an arterial line. Serum vials for analysis of cytokines were separated and kept deep frozen at -80°C until measurement. Another group of blood samples was used to analyze plasma FA composition with high-performance gas chromatography.

Inflammatory Response

During the treatment course, vital signs were recorded, including heart rate, blood pressure, respiration rate, body temperature, and other clinical parameters. White blood cell count and serum C-reactive protein (CRP) were also measured. The plasma level of interleukin (IL)-6 was quantified with an enzyme-linked immunosorbent assay (Jinmei Corp, Shanghai, China).

The ratio of systemic inflammatory response syndrome (SIRS) was recorded before and after PN in both groups. In 1992, SIRS was first defined by the American College of Chest Physicians/Society of Critical Care Medicine Consensus Conference. SIRS represents a hypermetabolic phase and dropout release of various proinflammatory factors. It typically has 2 or more of the 4 clinical findings: (1) white blood cell count > 12,000 cells/mm^sup 3^ or <4000 cells/mm^sup 3^ or >10% immature forms, (2) body temperature >38°C or <36°C, (3) heart rate >90 beats per minute, and (4) respiratory rate >20 breaths per minute or PaCO, <4.3 KPa. The SIRS score equals the number of clinical findings.14

Organ Function

Pulmonary function was measured by arterial blood gas to obtain the oxygenation index. The oxygenation index is the arterial oxygen pressure (PaO^sub 2^) divided by FiO^sub 2^. A SAP patient was considered to have acute respiratory distress syndrome (ARDS) if the PaO^sub 2^:FiO^sub 2^ ratio was <200 and there was a diffuse infiltrate (3 quadrants or more) on x-ray. In addition, there could be no evidence of a pulmonar)' capillary wedge pressure of > 2.4 kPa (18 mm Hg).15

Acute kidney injury is defined as an abrupt (within 48 hours) reduction in kidney function. The criteria include an absolute increase in serum creatinine of ≥0.3 mg/dL (≥26.4 µmol/L), a percentage increase in serum creatinine of ≥50% (1.5-fold from baseline), or reduced urine output (documented oliguria of <0.5 mL/kg/h for >6 hours). In SAP patients, the indications for continuous renal replacement therapy (CRRT) were acute kidney injury, metabolic acidosis, hyperkalemia, and so on.16

Outcome Measures

The primary outcome measures included infection morbidity, mortality, ICU stay, and the length of stay in the hospital.

Statistical Analysis

Values are presented as the mean ± SEM. Statistical analysis was performed by univariate ANOVA followed by a paired t test and Fisher exact test, only when the ANOVA was significant. SPSS 11.0 was used to perform the statistical analysis. P values <.05 were considered statistically significant.

Results

Patients

A total of 40 patients were included. The study was performed with 20 participants receiving the FO-supplemented fat emulsion (ω-3 FAs group) and 20 participants receiving the SO fat emulsion (control group). The PN was very well tolerated in all SAP patients in the 2 treatment groups. The distribution of age, gender, nutrition status, and the severity of SAP was comparable among the 2 groups (Table 1).

The Change of Plasma FA Profile

After 5 days of PN, there was a significant increase (P < .01) of eicosapentaenoic acid (EPA) content in the plasma in the FO-supplemented PN group but not in the control group (Figure 1).

Inflammatory Response

There were no significant differences in initial IL-6 levels prior to initiating PN. The level of IL-6 was reduced after PN in the ω-3 FAs group but slightly increased in the control group (Figure 2).

White blood cell count and CRP concentration before PN were not different between the 2 groups. The peak of CRP concentration was reached before initiating PN. The mean level was 165 mg/L (range, 60-264 mg/L) in the control group and 177 mg/L (range, 77-229 mg/L) in the ω-3 FAs group. After 5 days of PN, decreases in CRP concentration in both groups were statistically significant as compared with those on day 1 (P < .05). Moreover, reduction of CRP concentration was greater in the ω-3 FAs group than in the control group (P < .05; Figure 3). White blood cell counts were reduced from 14.16 ± 0.98 × 10^sup 9^/L to 13.22 ± 1.14 × 10^sup 9^/L in the control group, and from 15.09 ± 1.34 × 10^sup 9^/L to 12.49 ± 1.14 × 10^sup 9^/L in the ω-3 FAs group on day 6 after PN. However, these decreases were not significantly different between the 2 groups (Table 3).

The incidence of SIRS before PN were not different between the 2 groups (20/20 in both groups). After 5 days of PN, the incidence decreased significantly in both groups, with a pronounced decrease in the ω-3 FAs group but without a statistically significant difference compared with the control group (16/20 vs 11/20; Table 4).

Organ Dysfunction

The incidence of ARDS before PN in the 2 groups, which were measured to elucidate respiratory dysfunction in the acute phase of SAP, were not different between the 2 groups (14/20 in the ω-3 FAs group vs 13/20 in the control group). However, there were significant decreases in respiratory morbidity in both groups after PN infusion, and these decreases were not significantly different between the 2 groups. In addition, oxygenation indexes increased significantly in both groups from day 1 to day 6. We observed the favorable effect of FO on the lung, as documented by a significantly greater improvement of the oxygenation index after 5 days of FO-SO emulsion (P < .05; Table 4).

The morbidity of acute renal injury in SAP patients of the 2 groups was not significantly different (3/20 in the control group vs 2/20 in the ω-3 FAs group). However, the number of days of CRRT in the ω-3 FAs group was less than that in the control group (18 ± 2.3 days vs 26 ± 3.4 clays, P < .05; Table 4).

Outcome Measurements

All SAP patients were followed up for 1 month after discharge. In the control group, there were 5 patients with infectious complications (2 abscesses, 2 pneumonia, I wound infection), whereas 3 patients with infection (1 abscess, 1 pneumonia, 1 wound infection) were observed in the ω-3 FAs group.

There were no deaths in the ω-3 FAs group, whereas 2 patients died in the control group: 1 from multiple organ failure as a consequence of severe abdominal infection and the other from abdominal compartment syndrome after abdominal hemorrhage. All the other patients were transferred to general wards after ICU discharge.

The average ICU stay (21.4 ± 4.2 days in the ?-3 FAs group vs 27.5 ± 5.6 days in the control group) and hospital stay (65.2 ± 7.3 days in the ?-3 FAs group vs 70.5 ± 9.1 days in the control group) were not significantly different between the 2 groups.

Discussion

In the cascade of inflammatory response in the acute phase of SAP, there are treatment options for patients with SIRS or sepsis complicated by multiple organ failure. PN supplemented with ω-3 FAs is one of the strategies. ω-3 FAs in PN could compete with ω-6 FAs to change the composition of FAs in plasma. EPA, an ω-3 PUFA derived from FO, can rapidly incorporate into cell membrane phospholipids, replacing arachidonic acid as a substrate and converting into prostaglandin 3 (PG3) and leukotriene 5 (LT5) series.17 In the present study, we investigated the changes of plasma FA compositions in SAP patients and found an increased EPA concentration after 5 days of PN supplemented with ω-3 FAs.18,19 While ω-6 FAs have been demonstrated to enhance the inflammatory response and suppress host immunity through the production of potent inflammatory mediates such as prostaglandin 2 (PG2) and leukotriene 4 (LT4) series, ω-3 FAs are supposed to suppress inflammation and ameliorate the course of infection by reduction of proinflammatory eicosanoids and cytokines.5,7

De Caterina et al20 demonstrated that consumption of EPA decreased IL-6 and IL-8 in response to IL-1, tumor necrosis factor, or bacterial endotoxin. SAP can lead to a profound systemic inflammatory response. The hallmark of this response was activation of the acute phase through cytokines such as IL-6. In the ω-3 FAs group, we did find the decreased IL-6 level after PN emulsion, indicating a diminished inflammatory response. However, IL-6 levels increased in the control group. In theory, the modulation of eicosanoid and cytokine biology by the ω-3 FAs provides an intervention strategy for reducing the hyperinflammatory response so that eicosanoids and cytokines express beneficial effects rather than potentially damaging properties in SAP patients.21,22 Similar results were also observed in other reports.8,23,24

The question might be raised as to whether this treatment provided beneficial outcomes in SAP patients. Forty SAP patients were included, and 20 patients receiving supplemental FO did exhibit a significantly faster reduction in CRP level and in SIRS in the initial phase of SAP. We presume this reduction of hyperinflammation in SAP patients was at least partly due to the increase of fewer inflammatory mediators such as EPA in plasma and attenuation of IL-6. It can be deduced that ω-3 FAs were antiinflammatory lipids in SAP. Addition of FO may reduce the massive augmentation of the inflammatory response by ω-6 FA-based lipid emulsions and thereby preserve the inflammatory capacity and avoid substantial damage to organ function in SAP.

There was a greater increase in the oxygenation index after treatment with PN supplemented with ω-3 FAs treatment, showing that the respiratory function was improved in the ω-3 FAs group. In addition, the CRRT time was shorter in SAP patients with acute renal failure after ω-3 FAs supplementation compared with that after ω-6 FAs treatment, which indicated less damage to renal function. Although there were no significant differences of infection rate, ICU days, or length of hospital stay between the 2 groups, the trend of better clinical outcome with ω-3 FAs in SAP patients was observed in the present study.

In conclusion, our study shows that attenuation of the hyperinflammatory response can be obtained by ω-3 FAs supplementation, which changes plasma EPA concentration and decreases proinflammatory cytokines in severe acute pancreatitis. This, together with decreased SIRS ratio and improved respiratory and renal function, suggests that the systemic inflammatory response and other organ injury are attenuated, ω-3 FAs supplementation may thus be another tool for optimizing therapy in SAP that requires further experimental studies and clinical trials.