- Email: [email protected]
Effect of intralipid® on some immunological parameters and leukocyte functions in patients with esophageal and gastric cancer
CLINICAL NUTRITION
(1985) 4: 229-234
Effect of Intralipid* on Some Immunological Parameters and Leukocyte Functions in Patients with Esophageal and Gastric Cancer P.Dionigi,
R. Dionigi, U. Prati, F. Pavesi*, V. Jemos and S. Nazari
di Chirurgia, Sezione di Patologia Chirurgica, UniversitP di Pavia and *Servizio Analisi Chimico-Cliniche, Policlinico S. Matteo, IRCCS, 27100 Pavia, Italy (Reprints requests to P.D.)
Dipartimento
ABSTRACT This study has been undertaken to investigate if the intravenous (i.v.) infusion of fat emulsions may be associated with impairment of some immunological functions thus increasing the risk of septic complications. Fifteen malnourished patients with advanced gastric or esophageal cancer received for 2 weeks preoperatively and 1 week after surgery an isocaloric and isonitrogenous TPN treatment with Intralipid’” (group A: n = 8) or glucose alone (group B: n = 7) as energy substrate. Cluster analysis of 11 nutritional parameters and some tests of the humoral and cellular immunity (IgG, IgM, C3c, Factor B; polymorphonuclear (PMN) cells, total lymphocytes, T and B lymphocyte counts; ‘in vitro’ PMN chemotaxis, adherence to nylon fibers, phagocytosis of latex particles) were sequentially determined. The incidence and severity of post-operative infections were investigated and a ‘sepsis score’ was calculated for each patient. Preand postoperative TPN were not associated with an improvement of the nutritional status. The humoral and cellular immune parameters showed the same behaviour in patients receiving Intralipid”” and in controls. The chemotactic activity of PMN cells was constantly normal, granulocyte adherence fluctuated below the normality range in controls, whereas phagocytosis of latex was similar in both groups. Post-operative infectious episodes were less severe in patients receiving Intralipid. ” Our results do not confirm that Intralipid’” adversely affects some aspects of the humoral and cellular immune response. INTRODUCTION Undernutrition is frequently observed in patients with cancer of the upper gastrointestinal tract, and it plays a major role in increasing the risk of septic complications in the post-operative period [ 1,2,3]. Non specific abnormalities of the immune response have been observed in cancer patients and are associated with increased susceptibility to infections [4]. Clinical studies also indicate that infection itself may be a critical factor in precipitating acute nutritional deficiencies [5]. When oral alimentation is not possible, total parenteral nutrition (TPN) by means of the infusion of glucose, lipids and aminoacids is used for nutritional repletion and support in surgical patients before and after operation. Experimental and clinical studies have recently suggested that some mechanisms of the immune reactivity may be impaired during i.v. infusion of fat emulsions increasing the risk of infectious episodes [6, 7, 8, 91. On the contrary, other reports support evidence that addition of Intralipidm to the i.v. regimens does not adversely alter immune functions [lo, 11, 121 or may even improve them [ 131. 229
This prospective study has been undertaken in an of neoplastic patients to homogeneous group investigate whether Intralipid” can interfere with different aspects of the cellular and humoral immunological response before and after the surgical procedure.
PATIENTS
AND
METHODS
Patients
Twenty undernourished patients (weight loss > lo”,, of their usual body weight in the last 6 months) with a clinical diagnosis of advanced gastric or esophageal cancer were randomly allocated at admission into 2 groups (group A: TPN with IntralipiP; group B: TPN without Intralipid,“‘). Five patients were excluded from the study: 2 were found to be afflicted by benign lesions, and 3 needed more than 500mls of blood in the perioperative period. It has been demonstrated that blood loss is a possible cofactor of post-operative immunodepression [ 141. All the patients received a 24 h TPN regimen through a central vein providing the same amount of calories (50.6i8.5 kcal kg-‘day ‘) and nitrogen
230
EFFFX’I‘ OF IN’I’KI’~.IPII)
ON SORlE
I.\l.\lL~SOl.O~;I~:~\l.
(0.26 + 0.04 gN kg - 1 day-‘) for 2 weeks preoperatively and 1 week after surgery. In group A (n = 8; m/f =7/l; mean age 60 + 11 years) the 49:/h of the caloric supply was given as Intralipidfi’ and 51:~~ as glucose (52.11 i6.79 kcalkg-’ day-l), whereas patients in group B (n = 7; m/f= 3/4; mean age 65+ 12 years) received 1OOgl of calories as glucose (48.98 + 10.42 kcal kg-’ day-‘). Intralipidm was infused over a 16 h period allowing 8 h of night rest to avoid interference with the morning biochemical determinations. The total caloric and nitrogen supply was reduced on the day of operation and on the day after. Patients were allowed to drink only controlled amounts of water.
Investigations The nutritional status of the patients was evaluated at admission and twice a week before surgery (-12, - 9, -5 and -2 days) and after operation (+4, + 7, + 15 days) by means of cluster analysis of the following 11 parameters: y; usual body weight, arm total hemoglobin, serum muscle circumference, albumin, iron, transferrin, C3c, ceruloplasmin, retinol total lymphocytes and delayed binding protein, hypersensitivity response to four recall antigens and DNCB (only before surgery), according to the method previously described [ 151. In brief, cluster analysis identifies four clusters which represent different nutritional conditions. Patients belonging to cluster one are in good nutritional status, whereas patients belonging to cluster four are in the worst nutritional condition. Before starting TPN and at the same time as nutritional determinations the following immunoloserum were investigated: parameters gical concentration of IgG, IgM, Factor B (radial immunodiffusion technique, Behring Inst.), polyT and B morphonuclear (PMN) cell count, lymphocytes [16]. A chemotactic index was calculated: this is the sum of the leucotactic index of Maderazo and Woronick [ 171 and the maximum distance of migration (in pm) at which at least three cells per level are detectable. Granulocyte adherence to nylon fibers was determined following the method described by MacGregor and coworkers [18], modified for weight of the fibers employed (150mg of nylon) and for the time of incubation (18 min). PMN cell phagocytosis was studied in vitro employing the technique proposed by Hicks and Bennett [19] modified on the basis of our previous experience [20]; the method was simplified according to Wehinger
WI. Local effects effects of sepsis
of infection, and routine
pyrexia, laboratory
secondary data were
I’:\K.\ME’I’I~:KS
carefully recorded during the study. According to Elebute and Stoner [22] a score from 0 to 6 has been attributed to the parameters in proportion to their abnormalities. The sum of the scores gives the ‘sepsis score’ which reflects the overall severity of the infectious episodes; it is also correlated to the clinical outcome [23]. Tumor stage was evaluated following the UICC recommendations [24].
Statistical
analysis
The values sequentially determined within each group have been compared by means of analysis of variance (ANOVAR), and differences between groups have been evaluated by Student’s t test.
RESULTS Groups A and B were comparable in terms of preoperative nutritional status (Table l), mean tumor stage (3.5OkO.53 and 3.14f0.69) and daily amounts of calories and nitrogen supplied with TPN. Tumor was resectable (total or partial gastric resection, distal esophagectomy) in 75% of patients of group A and in 86% of patients of group B; palliative surgical procedure consisted in gastrostomy or jejunostomy. The duration of anesthesia and surgery were similar in groups A and B (196f98 vs 213+99min). Patients considered were malnourished at admission on the basis of their weight loss (group A: 16.4&9.39&; group B: 12 5+2.7oi,, nevertheless albumin levels (group A: ’ 365kO.44; group B: 3.41 kO.51) and cluster distributions (Table 1) were suggestive for a moderate degree of undernutrition. DHR was severely compromised in most of the patients (% hypo-anergic: group A: = 100%; group B=71%). Pre and postoperative TPN treatment was not associated with an improvement of the nutritional status when sequentially evaluated by means of cluster analysis. Table 1 shows that after surgery the nutritional condition of patients of group A was relatively stable, whereas it was more compromised in group B. At admission the basal determinations of the immune proteins except of IgM were similar in both groups and they did not show any significant variation for the whole period of study (Table 2). C3c mean values were high and remained above the Peripheral PMN cell counts normality range. increased significantly after surgery only in patients receiving fat emulsion. At the beginning of the study and during TPN total lymphocyte counts were markedly reduced in both groups. This reduction, which was statistically significant in group B, is
modifications of the nutritional Table 1 Sequential (OF’) indicates the surgical procedure
status
evaluated
by cluster
analysis.
Values
231
YIII‘KI I‘IOY
(:l.INI(:.‘\l.
are means
of clusters
k SD.
Weeks of TPN
2nd
Basal cluster Group Group
A B
1.9kO.35 2.OkO.5
Significance
of differences
2.50f0.8 2.43 k 0.8 between
2.3kl.l 2.0f0.8
2.38kO.9 2.14kO.7
groups:
3rd
2.0*0 2.4rfIO.5
2.OkO.8 2.2kO.7
Final cluster 2.1 kO.4 2.3kO.8
2.1kO.4 3.0+0.7=
“P < 0.05.
modifications of the immunological parameters. Table 2 Sequential subjects. Values are means+ SD. (OF) indicates the surgical procedure
Values
in brackets
have been obtained
from 25 normal
Weeks of TPN
, Group
3
1st
Basal
2nd
3rd
Final
Anovar
(GE)
(1330*252mg?,,)
A B
Ig M (156+96mg”,)
A B
172.9k47.8 123.1 f 16.8a
176.lf47.1 128.4 + 24.5”
c3c (85 + 20 mguo)
A B
llO.lk28.8 131.Ok38.6
Factor B (2Of6mg”,,)
A B
PMN cells (365Ok 1895//d)
n.s.
1361+438 1337k279
1398f514 1404f323
ll.S.
178.1 k51.2 133.Ok27.8
176.7 k50.9 141.1f32.7
173.0f75.2 138.3k26.5
n.s. n.s.
125.2 + 53.9 130.6f21.3
114.9k30.8 128.2? 17.2
136.1 f31.4 127.8 k 53.9
123.9k22.0 111.9f44.5
ll.S.
20.4* 10.7 22.4k9.9
20.5 k 4.7 20.9 + 4.5
22.1 k5.7 20.4k5.2
A B
3688 + 1025 5137k 1716
3611f1556 5753 k 5769
4000 * 1405 4165+ 1032
7425 f 3535 4824 k 2926
4418+2184 3588+815
PCO.005
Tot. lymphocytes (1773 + 834//d)
A B
1475*747 1375+354
951*441 983 + 480
671?203 888 * 449
1025k714 572 + 274
1002+740 572+352
n.s. r<:o.o5
T lymphocytes (1158f651/$)
A B
1089k581 902 + 296
633 f 276 775k416
546$- 166 646 f 260
696+411 447 + 256
B lymphocytes (195&71//d)
A B
k G
Significance
mainly
due
of differences
to a decrease
1336+510 1268k293
1258f318 1177k281
groups:
(“) Pi
of T lymphocytes,
25.9k7.2 22.9k7.1
69+25 70*35
77&55 69k33
lOOf 54+27
between
1381+509 1223f 196
82+66 59+75
25.6k7.0 25.8+ 11.9
652 + 425 472 k 299 81+65 39+29
n.s. ns. n.s. ILS.
Cl.S. C2.S. n.s. tl.S.
0.05
280
whereas
B lymphocytes are constantly low. The PMN cell chemotactic activity was normal in both groups with minor modifications after surgery in controls (Fig. 1). The granulocyte adherence to nylon fibers was normal and stable in patients receiving IntralipidR, but it fluctuated below the range of normality at the beginning of the TPN treatment and immediately after the surgical procedure in group B (Fig. 2). The in vitro phagocytosis of latex by neutrophils was initially high for all the patients when compared to the normal response (Fig. 3); it was not affected by TPN treatment and showed only a slight and transient increase which lasted a week after surgery and was similar in both groups.
1
r----
__---~p~~_________,
NEEKS
modifications of the chemotactic index Fig. 1 Sequential in patients receiving Intralipidn (group A= open columns) and without lipids (group B = dark columns). B = basal determinations. Arrow indicates the surgical procedure. Each column represents the mean f SD. Normal values from 25 normal subjects = 127 + 5 1.
El‘lW:I‘
OF
IK'I'K.4I.Il'II) OS
SOhlE
l\l\l1-SOl.O~;l~:.\l
T
I0 e
1
.y#
2
WEEKC
4 _I
Fig. 2 Sequential modifications of granulocyre adherence. Symbols as in Fig. 1. Normal values fiiom 2.5 normal subjects=85* 7”,,.
r---------
TV,.,----_
----
~
Fig. 3 Sequential modifications of phagocytosis of latex particles. Symbols as in Fig. 1. Normal values from 25 normal subjects = 35 + lSo/,,
A total of 13 post-operative septic complications were observed in 7 patients of group A, and 15 infectious episodes in 6 patients of group B. When the diagnosis of infection was made, patients were carefully monitored, and the ‘sepsis score’ was determined for each patient, showing a lower mean value in group A than group B (Table 3). This difference is statistically significant (P
DISCUSSION Controversial
Table 3
Group A Group B
results
concerning
the
relationships
1)\K.\\IE:'IEKS
between fat emulsion infusion and the immunological response have been recently published. Type and dose of the lipid substrate seem to interfere in different ways with the activity of granulocytes, R.E.S. circulating cells (monocytes) or fixed cells (macrophages). Impaired chemotaxis, lipid particle the engulfment in cytoplasm, dose-dependent decrease of the bactericidal capacity and nitroblue tetrazolium dye reduction have been described in granulocytes incubated “in vitro” with IntralipidVL or obtained from normal volunteers and patients receiving i.v. lipid emulsions 16, 7, 8, 251. Palmblad [12] did not find any abnormal “in vitro” migration, bactericidal function and chemiluminescence of neutrophil granulocytes in patients with Crohn’s disease receiving TPN with Intralipid”’ for a long period. A direct inhibitory effect of 20”,, Intralipid” on chemotaxis of monocytes has been observed by Fraser [26]; on the contrary, Wiernik has shown that “in vitro” preincubation of monocytes with the same lipid emulsion improves the ability of these cells to migrate towards a chemoattractor and to phagocytize yeast particles [25]. Moreover, Intralipid”’ has been reported to depress the “in vitro” response of human lymphocytes stimulated by PHA. The plasma of patients receiving fat emulsions shows the same inhibitory effect [27]. Employing soybean or safflower oil emulsions other authors describe an improvement of the blastogenic response of lymphocytes [ 10, 13, 261. Although immunodepression from both anesthesia and surgery have been amply documented, the relarive importance of each is virtually unknown [4,14]. Our results show that T and B lymphocytes, chemotaxis and granulocyte adherence are depressed after surgery only in the controls. The present study was undertaken in a homogeneous population of patients, considered to be at risk of septic complications. TPN was associated in group A with a high daily dose of lipids, and the treatment was maintained for 3 weeks. A relatively high amount of energy was infused in both groups to stress any possible effect of the different substrates on the immune response. TPN treatment was able to maintain a stable nutritional status, but did not
Incidence, onset, severity and duration of post-operative
septic complications.
Values are mean k SD
Incidence of infections (46 septic pts.)
Post-op. onset of infections (days)
Severity of infections (sepsis score)
Duration of sepsis (days)
Post-operative hospital stay (days)
87.5 85.7
4.1+2.0 3.Okl.O
8.9+_1.8, 15.7+3.0
14.Ok7.3 15.0&8.5
2l.Ok7.0 23.7 k9.6
Significance of differences between groups: “P< 0.001
CL.INICAL
induce any improvement. This is probably due to the fact that patients were moderately malnourished and the total amount of nitrogen supplied was just cover their daily requirements. sufficient to Nevertheless, no metabolic complications have been observed. IgG, IgM and Factor B remained within the range of normality and were not influenced by TPN treatment and Intralipid@ infusion. High levels of C3c have been recorded confirming previous studies in cancer patients showing that high concentrations of this complement component correlate well with the stage of the tumor [28]. Low peripheral lymphocyte counts are frequent in malnourished individuals with neoplasias, and seem to be reversed by nutritional repletion [29]. The lack of nutritional benefit that we observed was associated to a progressive reduction of total and T lymphocytes; nevertheless, this effect was more marked in patients receiving glucose as the only caloric substrate. No significant differences were detectable between the two groups with respect to the number of PMN cells and their chemotactic index, suggesting that the substrate composition should not be a critical determinant. In a previous work on 10 normal volunteers we observed a transient but significant fall of granulocyte adherence to nylon fibers during a 6 h i.v. administration of Intralip@ [30]. This effect reversed after the interruption of the infusion and the granulocyte adherence returned to normality within 6 h. In the present study the granulocyte adherence
NUTRITION
233
test was performed 8-Q h after the suspension of the lipid administration, and this may account for the absence of modification in this group. Surprisingly, more important alterations are presented by patients of group B at the beginning and at the end of TPN, and in relation to the surgical procedure. The phagocytic activity of PMN cells was not impaired in the patients who received Intralipid’“‘, and was similar to the control’s one. Our results are in agreement with the observations of Palmblad [12] but they are in contrast with other recent contributions [6, 251. In this study we have investigated some of the host defence mechanisms which participate in the bactericidal activity, such as the chemotactic response of phagocytic cells, their adherence to the foreign body and their phagocytic capacity. In general not significant differences have been observed between the two groups all over the study, even if patients receiving Intralipid ‘
REFERENCES Ill Diongi P, Nazari S, Bonoldi A P et al 1982 Nutritional assessment and surgical infections in patients with gastric cancer or peptic ulcer. Journal of Parenteral and Enteral Nutrition 6: 128-133 [21 Dionigi R, Genes F, Bonera A et al 1979 Nutrition and infection. Journal of Parenteral and Enteral Nutrition 3: 62-68 [31 Alexander J W 1975 Nutrition and surgical infections. In: American College of Surgeons, Battinger W (eds). Manual of surgical nutrition. WS Saunders Company, Philadelphia, pp 386-395 141 Dionigi R, Campani M 1981 Nutritional and immunological abnormalities in malignant disease. Acta Chirurgica Scandinavica, Suppl 507: 435-474 [51 Suskind R M, Sirisinha S, Edelman R et al 1978 Host defenses in protein-calorie malnutrition. In: Burke J F (ed). The infection prone hospital patient. Little Brown and Co, Boston, pp 121-142 161 Jarstrand C, Berghem L, Lahnborg G 1978 Human granulocyte and reticuloendothelial system function during Intralipid infusion. Journal of Parenteral and Enteral Nutrition 2: 663-670 171 Nordenstrom J, Jarstrand C, Wiemik A 1979
[8]
[91
[lOI
1111
[121
Decreased chemotactic and random migration of leukocytes during Intralipid infusion. American Journal of Clinical Nutrition 32: 2416-2422 Fischer G W, Hunter K W, Wilson S R, Mease A D 1980 Diminished bacterial defences with Intralipid. Lancet ii, 819-820 Loo L S, Tang J P, Kohl S 1982 Inhibition of cellular cytotoxicity of leukocytes for Herpes Simplex virusinfected cells in vitro and in vivo by Intralipid. Journal of Infectious Diseases 146: 6470 Helms R A, Herrod H G, Burckart G J et al 1983 E-rosette formation, total T-cells and lymphocyte transformation in infants receiving intravenous safflower oil emulsion. Journal of Parenteral and Enteral Nutrition 7: 541-545 Ota D M, Jessup J M, Babcock G F et al 1984 Immune function during soybean oil emulsion infusion. Journal of Parenteral and Enteral Nutrition 8: 84 Palmblad J, Brostrom 0, Lahnborg G et al 1982 Neutrophil functions during total parenteral nutrition and Intralipid infusion. American Journal of Clinical Nutrition 35: 1430-1436
234
EFFECT
OF
Ih’TR.4LIPID
ON
SOME
IMMC~NOLO(;IC,4L
[13] Ota D M, Copeland III E M, Corriere J N et al 1978 The effects of a loo/, soybean oil emulsion on lymphocyte transformation. Journal of Parenteral and Enteral Nutrition 2: 112-l 15 1141 Hubert A V. Lee E T, Hersh E M et al 1983 Effects of surgery, anesthesia and intraoperative blood loss on immunocompetence. Journal of Surgical Research 15: 399-403 [I51 Nazari S, Comincioli V, Dionigi R et al 1981 Cluster analysis of nutritional and immunological indicators for identification of high risk surgical patients. Journal of Parenteral and Enteral Nutrition 5: 307-316 [I61 Jondal M, Holm G, Wigzell H 1972 Surface markers on human T and B lymphocytes. I. A large population of lymphocytes forming non immune rosettes with sheep red blood cells. Journal of Experimental Medicine 136: 207-215 [I71 Maderazo E G. Woronick C L 1979 A modified micropore filter assay of human granulocyte leukotaxis. In: Gallin J I, Quie P G (eds), Leukocyte chemotaxis. Raven Press, New York, pp 43-54 [W MacGregor R R, Spagnuolo P J, Lentnek A L 1974 Inhibition of granulocyte adherence by ethanol, prednisone and aspirin measured with an assay system. New England Journal of Medicine 291: 642-646 t191 Hicks R G, Bennett J M 1971 An improved cytochemical method for nitroblue tetrazolium reduction by neutrophils. American Journal of Medical Technology 37: 226-229 [201 Gnes F, Galbiati A, Dominioni L, et al 1978 Funzionalita dei granulociti neutrotili in pazienti neoplastici sottoposti ad intervento chirurgico e chemioterapia. Bollettino Societa Medico Chirurgico 92, 157-163 Submission date: 2 May 1985. Accepted after Revision: 18 July 1985
PARAMETERS
WI Wehinger H, Hofacker M 1976 Latex phagocytosis by
polymorphonuclear leukocytes. European Journal of Pediatrics 123: 125-132 [221 Elebute E A, Stoner H B 1983 The grading of sepsis. British Journal of Surgery 70: 29-31 1231 Dionigi R, Dominioni L, Jemos V, et al 1985 Sepsis score and complement factor B for monitoring severely septic surgical patients and for predicting their survival. European Surgical Research 17: 320-329 [241 UICC 1979 TNM, Classification of malignant tumors. Minerva Medica, Torino c251 Wiemik A, Jarstrand C, Julander I 1983 The effect of Intralipid on mononuclear and polymorphonuclear phagocytes. American Journal of Clinical Nutrition 37: 256-26 1 WI Fraser I, Neoptolemos J, Woods I’, et al 1983 The effect of Intralipid on human lymphocyte and monocyte function. Clinical Nutrition 2: 37-40 v71 De Simone C, Ferrari M, Meli D, et al 1982 Reversibility by L-carnitine of immunosuppression induced by an emulsion of soybean oil, glycerol and egg lecithin. Arzneim Forsch/Drug Research 32: 14851488 WI Verhaegen H, de Cock W, de Cree J, et al 1976 Increase of serum complement levels in cancer patients with progressive tumors. Cancer 38: 1608-1613 P91 Kahan B D 1981 Nutrition and host defense mechanisms. Surgical Clinics of North America 61: 557-560 [301 Prati U, Pecis C, Tibaldeschi C, et al 1982 Studio della adesivita e chemiotassi leucocitaria durante infusione di Intralipid. Chirurgia Italiana 34: 42H31
(1985) 4: 229-234
Effect of Intralipid* on Some Immunological Parameters and Leukocyte Functions in Patients with Esophageal and Gastric Cancer P.Dionigi,
R. Dionigi, U. Prati, F. Pavesi*, V. Jemos and S. Nazari
di Chirurgia, Sezione di Patologia Chirurgica, UniversitP di Pavia and *Servizio Analisi Chimico-Cliniche, Policlinico S. Matteo, IRCCS, 27100 Pavia, Italy (Reprints requests to P.D.)
Dipartimento
ABSTRACT This study has been undertaken to investigate if the intravenous (i.v.) infusion of fat emulsions may be associated with impairment of some immunological functions thus increasing the risk of septic complications. Fifteen malnourished patients with advanced gastric or esophageal cancer received for 2 weeks preoperatively and 1 week after surgery an isocaloric and isonitrogenous TPN treatment with Intralipid’” (group A: n = 8) or glucose alone (group B: n = 7) as energy substrate. Cluster analysis of 11 nutritional parameters and some tests of the humoral and cellular immunity (IgG, IgM, C3c, Factor B; polymorphonuclear (PMN) cells, total lymphocytes, T and B lymphocyte counts; ‘in vitro’ PMN chemotaxis, adherence to nylon fibers, phagocytosis of latex particles) were sequentially determined. The incidence and severity of post-operative infections were investigated and a ‘sepsis score’ was calculated for each patient. Preand postoperative TPN were not associated with an improvement of the nutritional status. The humoral and cellular immune parameters showed the same behaviour in patients receiving Intralipid”” and in controls. The chemotactic activity of PMN cells was constantly normal, granulocyte adherence fluctuated below the normality range in controls, whereas phagocytosis of latex was similar in both groups. Post-operative infectious episodes were less severe in patients receiving Intralipid. ” Our results do not confirm that Intralipid’” adversely affects some aspects of the humoral and cellular immune response. INTRODUCTION Undernutrition is frequently observed in patients with cancer of the upper gastrointestinal tract, and it plays a major role in increasing the risk of septic complications in the post-operative period [ 1,2,3]. Non specific abnormalities of the immune response have been observed in cancer patients and are associated with increased susceptibility to infections [4]. Clinical studies also indicate that infection itself may be a critical factor in precipitating acute nutritional deficiencies [5]. When oral alimentation is not possible, total parenteral nutrition (TPN) by means of the infusion of glucose, lipids and aminoacids is used for nutritional repletion and support in surgical patients before and after operation. Experimental and clinical studies have recently suggested that some mechanisms of the immune reactivity may be impaired during i.v. infusion of fat emulsions increasing the risk of infectious episodes [6, 7, 8, 91. On the contrary, other reports support evidence that addition of Intralipidm to the i.v. regimens does not adversely alter immune functions [lo, 11, 121 or may even improve them [ 131. 229
This prospective study has been undertaken in an of neoplastic patients to homogeneous group investigate whether Intralipid” can interfere with different aspects of the cellular and humoral immunological response before and after the surgical procedure.
PATIENTS
AND
METHODS
Patients
Twenty undernourished patients (weight loss > lo”,, of their usual body weight in the last 6 months) with a clinical diagnosis of advanced gastric or esophageal cancer were randomly allocated at admission into 2 groups (group A: TPN with IntralipiP; group B: TPN without Intralipid,“‘). Five patients were excluded from the study: 2 were found to be afflicted by benign lesions, and 3 needed more than 500mls of blood in the perioperative period. It has been demonstrated that blood loss is a possible cofactor of post-operative immunodepression [ 141. All the patients received a 24 h TPN regimen through a central vein providing the same amount of calories (50.6i8.5 kcal kg-‘day ‘) and nitrogen
230
EFFFX’I‘ OF IN’I’KI’~.IPII)
ON SORlE
I.\l.\lL~SOl.O~;I~:~\l.
(0.26 + 0.04 gN kg - 1 day-‘) for 2 weeks preoperatively and 1 week after surgery. In group A (n = 8; m/f =7/l; mean age 60 + 11 years) the 49:/h of the caloric supply was given as Intralipidfi’ and 51:~~ as glucose (52.11 i6.79 kcalkg-’ day-l), whereas patients in group B (n = 7; m/f= 3/4; mean age 65+ 12 years) received 1OOgl of calories as glucose (48.98 + 10.42 kcal kg-’ day-‘). Intralipidm was infused over a 16 h period allowing 8 h of night rest to avoid interference with the morning biochemical determinations. The total caloric and nitrogen supply was reduced on the day of operation and on the day after. Patients were allowed to drink only controlled amounts of water.
Investigations The nutritional status of the patients was evaluated at admission and twice a week before surgery (-12, - 9, -5 and -2 days) and after operation (+4, + 7, + 15 days) by means of cluster analysis of the following 11 parameters: y; usual body weight, arm total hemoglobin, serum muscle circumference, albumin, iron, transferrin, C3c, ceruloplasmin, retinol total lymphocytes and delayed binding protein, hypersensitivity response to four recall antigens and DNCB (only before surgery), according to the method previously described [ 151. In brief, cluster analysis identifies four clusters which represent different nutritional conditions. Patients belonging to cluster one are in good nutritional status, whereas patients belonging to cluster four are in the worst nutritional condition. Before starting TPN and at the same time as nutritional determinations the following immunoloserum were investigated: parameters gical concentration of IgG, IgM, Factor B (radial immunodiffusion technique, Behring Inst.), polyT and B morphonuclear (PMN) cell count, lymphocytes [16]. A chemotactic index was calculated: this is the sum of the leucotactic index of Maderazo and Woronick [ 171 and the maximum distance of migration (in pm) at which at least three cells per level are detectable. Granulocyte adherence to nylon fibers was determined following the method described by MacGregor and coworkers [18], modified for weight of the fibers employed (150mg of nylon) and for the time of incubation (18 min). PMN cell phagocytosis was studied in vitro employing the technique proposed by Hicks and Bennett [19] modified on the basis of our previous experience [20]; the method was simplified according to Wehinger
WI. Local effects effects of sepsis
of infection, and routine
pyrexia, laboratory
secondary data were
I’:\K.\ME’I’I~:KS
carefully recorded during the study. According to Elebute and Stoner [22] a score from 0 to 6 has been attributed to the parameters in proportion to their abnormalities. The sum of the scores gives the ‘sepsis score’ which reflects the overall severity of the infectious episodes; it is also correlated to the clinical outcome [23]. Tumor stage was evaluated following the UICC recommendations [24].
Statistical
analysis
The values sequentially determined within each group have been compared by means of analysis of variance (ANOVAR), and differences between groups have been evaluated by Student’s t test.
RESULTS Groups A and B were comparable in terms of preoperative nutritional status (Table l), mean tumor stage (3.5OkO.53 and 3.14f0.69) and daily amounts of calories and nitrogen supplied with TPN. Tumor was resectable (total or partial gastric resection, distal esophagectomy) in 75% of patients of group A and in 86% of patients of group B; palliative surgical procedure consisted in gastrostomy or jejunostomy. The duration of anesthesia and surgery were similar in groups A and B (196f98 vs 213+99min). Patients considered were malnourished at admission on the basis of their weight loss (group A: 16.4&9.39&; group B: 12 5+2.7oi,, nevertheless albumin levels (group A: ’ 365kO.44; group B: 3.41 kO.51) and cluster distributions (Table 1) were suggestive for a moderate degree of undernutrition. DHR was severely compromised in most of the patients (% hypo-anergic: group A: = 100%; group B=71%). Pre and postoperative TPN treatment was not associated with an improvement of the nutritional status when sequentially evaluated by means of cluster analysis. Table 1 shows that after surgery the nutritional condition of patients of group A was relatively stable, whereas it was more compromised in group B. At admission the basal determinations of the immune proteins except of IgM were similar in both groups and they did not show any significant variation for the whole period of study (Table 2). C3c mean values were high and remained above the Peripheral PMN cell counts normality range. increased significantly after surgery only in patients receiving fat emulsion. At the beginning of the study and during TPN total lymphocyte counts were markedly reduced in both groups. This reduction, which was statistically significant in group B, is
modifications of the nutritional Table 1 Sequential (OF’) indicates the surgical procedure
status
evaluated
by cluster
analysis.
Values
231
YIII‘KI I‘IOY
(:l.INI(:.‘\l.
are means
of clusters
k SD.
Weeks of TPN
2nd
Basal cluster Group Group
A B
1.9kO.35 2.OkO.5
Significance
of differences
2.50f0.8 2.43 k 0.8 between
2.3kl.l 2.0f0.8
2.38kO.9 2.14kO.7
groups:
3rd
2.0*0 2.4rfIO.5
2.OkO.8 2.2kO.7
Final cluster 2.1 kO.4 2.3kO.8
2.1kO.4 3.0+0.7=
“P < 0.05.
modifications of the immunological parameters. Table 2 Sequential subjects. Values are means+ SD. (OF) indicates the surgical procedure
Values
in brackets
have been obtained
from 25 normal
Weeks of TPN
, Group
3
1st
Basal
2nd
3rd
Final
Anovar
(GE)
(1330*252mg?,,)
A B
Ig M (156+96mg”,)
A B
172.9k47.8 123.1 f 16.8a
176.lf47.1 128.4 + 24.5”
c3c (85 + 20 mguo)
A B
llO.lk28.8 131.Ok38.6
Factor B (2Of6mg”,,)
A B
PMN cells (365Ok 1895//d)
n.s.
1361+438 1337k279
1398f514 1404f323
ll.S.
178.1 k51.2 133.Ok27.8
176.7 k50.9 141.1f32.7
173.0f75.2 138.3k26.5
n.s. n.s.
125.2 + 53.9 130.6f21.3
114.9k30.8 128.2? 17.2
136.1 f31.4 127.8 k 53.9
123.9k22.0 111.9f44.5
ll.S.
20.4* 10.7 22.4k9.9
20.5 k 4.7 20.9 + 4.5
22.1 k5.7 20.4k5.2
A B
3688 + 1025 5137k 1716
3611f1556 5753 k 5769
4000 * 1405 4165+ 1032
7425 f 3535 4824 k 2926
4418+2184 3588+815
PCO.005
Tot. lymphocytes (1773 + 834//d)
A B
1475*747 1375+354
951*441 983 + 480
671?203 888 * 449
1025k714 572 + 274
1002+740 572+352
n.s. r<:o.o5
T lymphocytes (1158f651/$)
A B
1089k581 902 + 296
633 f 276 775k416
546$- 166 646 f 260
696+411 447 + 256
B lymphocytes (195&71//d)
A B
k G
Significance
mainly
due
of differences
to a decrease
1336+510 1268k293
1258f318 1177k281
groups:
(“) Pi
of T lymphocytes,
25.9k7.2 22.9k7.1
69+25 70*35
77&55 69k33
lOOf 54+27
between
1381+509 1223f 196
82+66 59+75
25.6k7.0 25.8+ 11.9
652 + 425 472 k 299 81+65 39+29
n.s. ns. n.s. ILS.
Cl.S. C2.S. n.s. tl.S.
0.05
280
whereas
B lymphocytes are constantly low. The PMN cell chemotactic activity was normal in both groups with minor modifications after surgery in controls (Fig. 1). The granulocyte adherence to nylon fibers was normal and stable in patients receiving IntralipidR, but it fluctuated below the range of normality at the beginning of the TPN treatment and immediately after the surgical procedure in group B (Fig. 2). The in vitro phagocytosis of latex by neutrophils was initially high for all the patients when compared to the normal response (Fig. 3); it was not affected by TPN treatment and showed only a slight and transient increase which lasted a week after surgery and was similar in both groups.
1
r----
__---~p~~_________,
NEEKS
modifications of the chemotactic index Fig. 1 Sequential in patients receiving Intralipidn (group A= open columns) and without lipids (group B = dark columns). B = basal determinations. Arrow indicates the surgical procedure. Each column represents the mean f SD. Normal values from 25 normal subjects = 127 + 5 1.
El‘lW:I‘
OF
IK'I'K.4I.Il'II) OS
SOhlE
l\l\l1-SOl.O~;l~:.\l
T
I0 e
1
.y#
2
WEEKC
4 _I
Fig. 2 Sequential modifications of granulocyre adherence. Symbols as in Fig. 1. Normal values fiiom 2.5 normal subjects=85* 7”,,.
r---------
TV,.,----_
----
~
Fig. 3 Sequential modifications of phagocytosis of latex particles. Symbols as in Fig. 1. Normal values from 25 normal subjects = 35 + lSo/,,
A total of 13 post-operative septic complications were observed in 7 patients of group A, and 15 infectious episodes in 6 patients of group B. When the diagnosis of infection was made, patients were carefully monitored, and the ‘sepsis score’ was determined for each patient, showing a lower mean value in group A than group B (Table 3). This difference is statistically significant (P
DISCUSSION Controversial
Table 3
Group A Group B
results
concerning
the
relationships
1)\K.\\IE:'IEKS
between fat emulsion infusion and the immunological response have been recently published. Type and dose of the lipid substrate seem to interfere in different ways with the activity of granulocytes, R.E.S. circulating cells (monocytes) or fixed cells (macrophages). Impaired chemotaxis, lipid particle the engulfment in cytoplasm, dose-dependent decrease of the bactericidal capacity and nitroblue tetrazolium dye reduction have been described in granulocytes incubated “in vitro” with IntralipidVL or obtained from normal volunteers and patients receiving i.v. lipid emulsions 16, 7, 8, 251. Palmblad [12] did not find any abnormal “in vitro” migration, bactericidal function and chemiluminescence of neutrophil granulocytes in patients with Crohn’s disease receiving TPN with Intralipid”’ for a long period. A direct inhibitory effect of 20”,, Intralipid” on chemotaxis of monocytes has been observed by Fraser [26]; on the contrary, Wiernik has shown that “in vitro” preincubation of monocytes with the same lipid emulsion improves the ability of these cells to migrate towards a chemoattractor and to phagocytize yeast particles [25]. Moreover, Intralipid”’ has been reported to depress the “in vitro” response of human lymphocytes stimulated by PHA. The plasma of patients receiving fat emulsions shows the same inhibitory effect [27]. Employing soybean or safflower oil emulsions other authors describe an improvement of the blastogenic response of lymphocytes [ 10, 13, 261. Although immunodepression from both anesthesia and surgery have been amply documented, the relarive importance of each is virtually unknown [4,14]. Our results show that T and B lymphocytes, chemotaxis and granulocyte adherence are depressed after surgery only in the controls. The present study was undertaken in a homogeneous population of patients, considered to be at risk of septic complications. TPN was associated in group A with a high daily dose of lipids, and the treatment was maintained for 3 weeks. A relatively high amount of energy was infused in both groups to stress any possible effect of the different substrates on the immune response. TPN treatment was able to maintain a stable nutritional status, but did not
Incidence, onset, severity and duration of post-operative
septic complications.
Values are mean k SD
Incidence of infections (46 septic pts.)
Post-op. onset of infections (days)
Severity of infections (sepsis score)
Duration of sepsis (days)
Post-operative hospital stay (days)
87.5 85.7
4.1+2.0 3.Okl.O
8.9+_1.8, 15.7+3.0
14.Ok7.3 15.0&8.5
2l.Ok7.0 23.7 k9.6
Significance of differences between groups: “P< 0.001
CL.INICAL
induce any improvement. This is probably due to the fact that patients were moderately malnourished and the total amount of nitrogen supplied was just cover their daily requirements. sufficient to Nevertheless, no metabolic complications have been observed. IgG, IgM and Factor B remained within the range of normality and were not influenced by TPN treatment and Intralipid@ infusion. High levels of C3c have been recorded confirming previous studies in cancer patients showing that high concentrations of this complement component correlate well with the stage of the tumor [28]. Low peripheral lymphocyte counts are frequent in malnourished individuals with neoplasias, and seem to be reversed by nutritional repletion [29]. The lack of nutritional benefit that we observed was associated to a progressive reduction of total and T lymphocytes; nevertheless, this effect was more marked in patients receiving glucose as the only caloric substrate. No significant differences were detectable between the two groups with respect to the number of PMN cells and their chemotactic index, suggesting that the substrate composition should not be a critical determinant. In a previous work on 10 normal volunteers we observed a transient but significant fall of granulocyte adherence to nylon fibers during a 6 h i.v. administration of Intralip@ [30]. This effect reversed after the interruption of the infusion and the granulocyte adherence returned to normality within 6 h. In the present study the granulocyte adherence
NUTRITION
233
test was performed 8-Q h after the suspension of the lipid administration, and this may account for the absence of modification in this group. Surprisingly, more important alterations are presented by patients of group B at the beginning and at the end of TPN, and in relation to the surgical procedure. The phagocytic activity of PMN cells was not impaired in the patients who received Intralipid’“‘, and was similar to the control’s one. Our results are in agreement with the observations of Palmblad [12] but they are in contrast with other recent contributions [6, 251. In this study we have investigated some of the host defence mechanisms which participate in the bactericidal activity, such as the chemotactic response of phagocytic cells, their adherence to the foreign body and their phagocytic capacity. In general not significant differences have been observed between the two groups all over the study, even if patients receiving Intralipid ‘
REFERENCES Ill Diongi P, Nazari S, Bonoldi A P et al 1982 Nutritional assessment and surgical infections in patients with gastric cancer or peptic ulcer. Journal of Parenteral and Enteral Nutrition 6: 128-133 [21 Dionigi R, Genes F, Bonera A et al 1979 Nutrition and infection. Journal of Parenteral and Enteral Nutrition 3: 62-68 [31 Alexander J W 1975 Nutrition and surgical infections. In: American College of Surgeons, Battinger W (eds). Manual of surgical nutrition. WS Saunders Company, Philadelphia, pp 386-395 141 Dionigi R, Campani M 1981 Nutritional and immunological abnormalities in malignant disease. Acta Chirurgica Scandinavica, Suppl 507: 435-474 [51 Suskind R M, Sirisinha S, Edelman R et al 1978 Host defenses in protein-calorie malnutrition. In: Burke J F (ed). The infection prone hospital patient. Little Brown and Co, Boston, pp 121-142 161 Jarstrand C, Berghem L, Lahnborg G 1978 Human granulocyte and reticuloendothelial system function during Intralipid infusion. Journal of Parenteral and Enteral Nutrition 2: 663-670 171 Nordenstrom J, Jarstrand C, Wiemik A 1979
[8]
[91
[lOI
1111
[121
Decreased chemotactic and random migration of leukocytes during Intralipid infusion. American Journal of Clinical Nutrition 32: 2416-2422 Fischer G W, Hunter K W, Wilson S R, Mease A D 1980 Diminished bacterial defences with Intralipid. Lancet ii, 819-820 Loo L S, Tang J P, Kohl S 1982 Inhibition of cellular cytotoxicity of leukocytes for Herpes Simplex virusinfected cells in vitro and in vivo by Intralipid. Journal of Infectious Diseases 146: 6470 Helms R A, Herrod H G, Burckart G J et al 1983 E-rosette formation, total T-cells and lymphocyte transformation in infants receiving intravenous safflower oil emulsion. Journal of Parenteral and Enteral Nutrition 7: 541-545 Ota D M, Jessup J M, Babcock G F et al 1984 Immune function during soybean oil emulsion infusion. Journal of Parenteral and Enteral Nutrition 8: 84 Palmblad J, Brostrom 0, Lahnborg G et al 1982 Neutrophil functions during total parenteral nutrition and Intralipid infusion. American Journal of Clinical Nutrition 35: 1430-1436
234
EFFECT
OF
Ih’TR.4LIPID
ON
SOME
IMMC~NOLO(;IC,4L
[13] Ota D M, Copeland III E M, Corriere J N et al 1978 The effects of a loo/, soybean oil emulsion on lymphocyte transformation. Journal of Parenteral and Enteral Nutrition 2: 112-l 15 1141 Hubert A V. Lee E T, Hersh E M et al 1983 Effects of surgery, anesthesia and intraoperative blood loss on immunocompetence. Journal of Surgical Research 15: 399-403 [I51 Nazari S, Comincioli V, Dionigi R et al 1981 Cluster analysis of nutritional and immunological indicators for identification of high risk surgical patients. Journal of Parenteral and Enteral Nutrition 5: 307-316 [I61 Jondal M, Holm G, Wigzell H 1972 Surface markers on human T and B lymphocytes. I. A large population of lymphocytes forming non immune rosettes with sheep red blood cells. Journal of Experimental Medicine 136: 207-215 [I71 Maderazo E G. Woronick C L 1979 A modified micropore filter assay of human granulocyte leukotaxis. In: Gallin J I, Quie P G (eds), Leukocyte chemotaxis. Raven Press, New York, pp 43-54 [W MacGregor R R, Spagnuolo P J, Lentnek A L 1974 Inhibition of granulocyte adherence by ethanol, prednisone and aspirin measured with an assay system. New England Journal of Medicine 291: 642-646 t191 Hicks R G, Bennett J M 1971 An improved cytochemical method for nitroblue tetrazolium reduction by neutrophils. American Journal of Medical Technology 37: 226-229 [201 Gnes F, Galbiati A, Dominioni L, et al 1978 Funzionalita dei granulociti neutrotili in pazienti neoplastici sottoposti ad intervento chirurgico e chemioterapia. Bollettino Societa Medico Chirurgico 92, 157-163 Submission date: 2 May 1985. Accepted after Revision: 18 July 1985
PARAMETERS
WI Wehinger H, Hofacker M 1976 Latex phagocytosis by
polymorphonuclear leukocytes. European Journal of Pediatrics 123: 125-132 [221 Elebute E A, Stoner H B 1983 The grading of sepsis. British Journal of Surgery 70: 29-31 1231 Dionigi R, Dominioni L, Jemos V, et al 1985 Sepsis score and complement factor B for monitoring severely septic surgical patients and for predicting their survival. European Surgical Research 17: 320-329 [241 UICC 1979 TNM, Classification of malignant tumors. Minerva Medica, Torino c251 Wiemik A, Jarstrand C, Julander I 1983 The effect of Intralipid on mononuclear and polymorphonuclear phagocytes. American Journal of Clinical Nutrition 37: 256-26 1 WI Fraser I, Neoptolemos J, Woods I’, et al 1983 The effect of Intralipid on human lymphocyte and monocyte function. Clinical Nutrition 2: 37-40 v71 De Simone C, Ferrari M, Meli D, et al 1982 Reversibility by L-carnitine of immunosuppression induced by an emulsion of soybean oil, glycerol and egg lecithin. Arzneim Forsch/Drug Research 32: 14851488 WI Verhaegen H, de Cock W, de Cree J, et al 1976 Increase of serum complement levels in cancer patients with progressive tumors. Cancer 38: 1608-1613 P91 Kahan B D 1981 Nutrition and host defense mechanisms. Surgical Clinics of North America 61: 557-560 [301 Prati U, Pecis C, Tibaldeschi C, et al 1982 Studio della adesivita e chemiotassi leucocitaria durante infusione di Intralipid. Chirurgia Italiana 34: 42H31