Serum Concentrations Of Proinflammatory Cytokines Research Articles

P269 - Serum pro-inflammatory cytokine concentrations in patients with Crohn's disease and osteoporosis

s of the 4th Congress of ECCO the European Crohn’s and Colitis Organisation S117 of MMP-9 and MMP-26 positive neutrophils and stromal TIMP-1 and TIMP-3. Epithelial MMP-7 expression tended to be lower in post-treatment samples and could be in an important player in the mucosal wound healing response elicited by TNF-alphainhibitors. MMP-1 and MMP-10 do not seem to be important players in this process. P269 Serum pro-inflammatory cytokine concentrations in patients with Crohn’s disease and osteoporosis C. Zavos1 *, I. Koutroubakis1, M. Kamba2, M. Niniraki2, S. Kolyvaki2, I. Nerantzoulakis3, T. Voudouri1, E. Kouroumalis1. 1Department of Gastroenterology, University Hospital of Heraklion, Heraklion Crete, Greece, 2Department of Immunology, University Hospital of Heraklion, Heraklion Crete, Greece, 3Department of Radiology, University Hospital of Heraklion, Heraklion Crete, Greece Background: A high prevalence of low bone mineralisation (LBM) is observed in inflammatory bowel diseases (IBD), particularly in Crohn’s disease (CD). Chronic intestinal lesions of CD patients are associated with a Th1 type cytokine profile. The expression of pro-inflammatory cytokines in intestinal mucosa from CD patients is markedly enhanced. Likewise, some of these cytokines, particularly interleukin (IL)-6, were found to play a role in the pathogenesis of postmenopausal osteoporosis. Aims: To investigate the role of pro-inflammatory cytokines IL-1b, -2, -4, -6, -8, -10, interferon (IFN)-g and tumour necrosis factor (TNF)-a in CD patients with LBM, particularly in the cases with osteoporosis. Patients and Methods: Sixty-three consecutive CD patients were included in the study, of whom 18 (9 women, mean age: 31.7±7.2 years) had T-score > 1.0, considered as normal; 29 (14 women, mean age: 42.7±13.0 years) had T-score between 1.0 and 2.5, indicative of osteopenia; and 16 (7 women, mean age: 53.9±19.2 years) had T-score < 2.5, suggestive of osteoporosis. All patients underwent bone densitometry by dual energy X-ray absorptiometry (DXA) at the femoral neck and lumbar spine levels (L1-L4). The serum concentrations of the afore-mentioned pro-inflammatory cytokines were determined by multiplex technique. Results: No significant differences in mean cytokine concentrations between patients with normal T-score and those with T-score suggesting osteopenia were found. On the other hand, significant differences between patients without LBM and those with osteoporosis were noticed. As shown in Table 1, IL-2, -6, -8, and -10 were found to be significantly elevated in CD patients with osteoporosis compared to those without LBM. Conversely, IFN-g, TNF-a, IL-1b, and IL-4 serum levels were similar in the 2 study groups. Patients with osteoporosis had similar CDAI and CRP levels to those without LBM.

Increased macrophage colony-stimulating factor levels in patients with Graves’ disease

Previous studies have found markedly elevated serum concentrations of proinflammatory cytokines in patients with Graves' disease (GD). We investigated the role of macrophage colony-stimulating factor (M-CSF) in GD. We assayed concentrations of M-CSF in sera from 32 patients with GD (25 untreated; 7 receiving thiamazole therapy). We also studied 32 age-matched healthy subjects as controls. Relationships between serum M-CSF and both thyroid state and serum lipids were examined. Moreover, to examine the effect of thyroid hormone alone on serum M-CSF, T3 was administered orally to normal subjects. Serum concentrations of M-CSF in GD patients who were hyperthyroid were significantly increased compared with GD patients who were euthyroid (P < 0.05) and control subjects (P < 0.0001). Serum M-CSF concentrations correlated closely with T3 levels in patients (r = 0.51, P < 0.005). Serial measurement of five individual patients revealed that serum concentrations of M-CSF were significantly decreased (P < 0.05), reaching normal control values upon attainment of euthyroidism. Furthermore, oral T3 administered to 15 volunteers for 7 days produced significant increases in serum levels of M-CSF (P < 0.05). The close correlation between serum M-CSF and serum thyroid hormone levels suggests that high circulating levels of thyroid hormones may directly or indirectly potentiate the production of M-CSF in patients with GD.

Associations of circulating adiponectin with estradiol and monocyte chemotactic protein-1 in postmenopausal women

The aim of the present study was to clarify the association of serum adiponectin concentrations with serum 17beta-estradiol concentrations in pre-, peri-, and postmenopausal women. In addition, the associations of serum adiponectin with serum concentrations of proinflammatory and anti-inflammatory cytokines were examined in women during the menopausal transition. A total of 197 women were enrolled in this study: 33 premenopausal women, 80 perimenopausal women, and 84 postmenopausal women. Serum adiponectin concentration was measured by an enzyme-linked immunosorbent assay. Serum concentrations of the proinflammatory cytokines interleukin (IL)-1beta, IL-6, and tumor necrosis factor alpha, anti-inflammatory cytokine IL-10, and the chemokines IL-8, macrophage inflammatory protein-1beta and monocyte chemotactic protein-1 were measured by using a multiplexed human cytokine assay. Serum adiponectin concentration showed a significant negative correlation with serum estradiol concentration (r= -0.400, P=0.001) in postmenopausal women but not in pre- and perimenopausal women, and this correlation was significant after adjustment for age and body mass index. Serum adiponectin concentration also showed a significant negative correlation with serum monocyte chemotactic protein-1 concentration (r= -0.244, P=0.05) in postmenopausal women. An increase in adiponectin level due to a decrease in estradiol results in a reduction in monocyte chemotactic protein-1 level in postmenopausal women, suggesting that adiponectin may be associated with a protective role against insulin resistance and atherosclerosis, which occur in the postmenopausal stage.

Decreased Serum Paraoxonase-1 Activity during Intestinal Nematode (Nippostrongylus brasiliensis) Infection in Rats

Reduced paraoxonase-1 (PON1) activity has been observed in a number of pathological conditions; however, little is known about the effects of intestinal nematode infections, such as Nippostrongylus brasiliensis, on paraoxonase activity. We observed a significant reduction in serum paraoxonase and arylesterase activity after N. brasiliensis infection in Wistar rats from Day 6 until Day 12 post-infection (p.i.) for serum paraoxonase and from Day 3 until Day 24 p.i. for arylesterase. In addition, N. brasiliensis infection increased serum concentrations of pro-inflammatory cytokines (interleukin-1, interleukin-6, and tumor necrosis factor-alpha), with maximum concentrations observed on Day 9 p.i. These cytokines are known to inhibit the synthesis of hepatic PON1 mRNA. Thus, the observed reduction in PON1 activity during N. brasiliensis infection is likely associated with inflammatory reactions mounted against the parasites.

Dietary α-linolenic acid inhibits proinflammatory cytokine production by peripheral blood mononuclear cells in hypercholesterolemic subjects

Background:Atherosclerosis is a chronic inflammatory disease. We previously reported that a diet high in α-linolenic acid (ALA) reduces lipid and inflammatory cardiovascular disease risk factors in hypercholesterolemic subjects.Objective:The objective was to evaluate the effects of a diet high in ALA on serum proinflammatory cytokine concentrations and cytokine production by cultured peripheral blood mononuclear cells (PBMCs) from subjects fed the experimental diets.Design:A randomized, controlled, 3-diet, 3-period crossover study design was used. Hypercholesterolemic subjects (n= 23) were assigned to 3 experimental diets: a diet high in ALA (ALA diet; 6.5% of energy), a diet high in linoleic acid (LA diet; 12.6% of energy), and an average American diet (AAD) for 6 wk. Serum interleukin (IL)-6, IL-1β, and tumor necrosis factor-α (TNF-α) concentrations and the production of IL-6, IL-1β, and TNF-α by PBMCs were measured.Results:IL-6, IL-1β, and TNF-α production by PBMCs and serum TNF-α concentrations were lower (P< 0.05 and P< 0.08, respectively) with the ALA diet than with the LA diet or AAD. PBMC production of TNF-α was inversely correlated with ALA (r= −0.402, P= 0.07) and with eicosapentaenoic acid (r= −0.476, P= 0.03) concentrations in PBMC lipids with the ALA diet. Changes in serum ALA were inversely correlated with changes in TNF-α produced by PBMCs (r= −0.423, P< 0.05).Conclusions:Increased intakes of dietary ALA elicit antiinflammatory effects by inhibiting IL-6, IL-1β, and TNF-α production in cultured PBMCs. Changes in PBMC ALA and eicosapentaenoic acid (derived from dietary ALA) are associated with beneficial changes in TNF-α release. Thus, the cardioprotective effects of ALA are mediated in part by a reduction in the production of inflammatory cytokines.

Prednisone (P) prophylaxis decreases incidence and severity of lenalidomide (L) induced flare reaction (flr) in patients with relapse (rel) or refractory (ref) chronic lymphocytic leukemia

18512 Background: Flare reaction (flr) is a new side effect unique to CLL pts treated with immunomodulating agents both thalidomide (T) and L. It is characterized by tender swelling of disease involved lymph nodes (LN), spleen, rash, low-grade fever and/or increase in absolute lymphocyte counts (ALC). Underlying biologic process remains unknown though it is postulated to be mediated through activation of immune effector cells and/or change in serum concentration of pro-inflammatory cytokines. In a phase II clinical trial of L in rel/ref CLL pts we noted a high incidence of flr. Here we report treatment schema for flr and a new preventive approach employed in this study. Methods: A total of 35 pts are enrolled. No prophylaxis given to initial cohort (A). Pts were treated at onset of symptoms with (1) ibuprofen (Ib) 600 mg po q6 hours (2) oxycodone (oxy) 5–10 mg po prn q 6 hours. In subsequent cohort (B) we are investigating prophylactic use of P 20 mg PO QD starting day −1 × 5 days followed by 10 mg QD × 5 days. Results: All pts had rel or ref disease. Median age 63 (range 48–76) year. In cohort A (n = 29) overall incidence of flr was 79% (n = 23). Of these the severity was graded as I, II and III in 48%, 24%, and 7% respectively. There was no grade IV flr. Symptoms initiated within hours of intake of the 1st dose of L lasting for a median of 14 days (cycle 1 only). Pts were adequately managed with symptom resolution using Ib/oxy. Recurrence in cycle 2 was noted in 1 pt. Pain in LN was the most common complaint. Increase ALC noted in 2 pts. Serum cytokine analysis (n = 5) pre and post 7-days of L treatment showed significant increase in TNF-α, IL-10 and MIG. In cohort B (n = 6) 2 pts had flr (grade I) lasting ≤ 10 days. Conclusions: Prophylactic use of P results in decrease incidence, severity and duration of flr. There was no correlation with clinical response. Elevation of ALC can be part of the flr, pts response should be interpreted cautiously for disease progression. Our findings suggest that use of P may be an effective prophylactic maneuver for prevention of L induced flr. Updated results of cohort B will be presented at this meeting. [Table: see text]

Transient Leukocytopenia Associated with a Steep Surge of Pro-inflammatory Cytokines in a Patient with Severe Cardiogenic Pulmonary Edema

Leukocytopenia can be caused by depressed production, increased peripheral destruction, or excessive peripheral pooling. Leukocyte margination is one of the mechanisms responsible for excessive peripheral pooling. A reversible leukocyte margination is caused by an increase in pro-inflammatory cytokines. However, there are limited data for this phenomenon in clinical conditions. We describe a case of unexpected transient leukocytopenia after exchanging an extracorporeal membrane oxygenation (ECMO) system used to treat severe cardiogenic pulmonary edema. To assess the cause of the leukocytopenia, the serum concentrations of pro-inflammatory cytokines and selectins were measured. The concentrations of tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, and IL-8 were markedly, but transiently, elevated in relation to the leukocytopenia. The transient leukocytopenia with pulmonary margination appeared to be caused by a steep surge of pro-inflammatory cytokines stimulated by hypoxia/reoxygenation during the exchange of the ECMO system. This case may suggest the mechanisms responsible for leukocytopenia in the clinical entity referred to as "systemic inflammatory response syndrome"

Modulation of TNF-α Gene Expression by IFN-γ and Pamidronate in Murine Macrophages: Regulation by STAT1-Dependent Pathways

Aminobisphosphonates are drugs used in the treatment of hypercalcemia, Paget's disease, osteoporosis, and malignancy. Some patients treated with aminobisphosphonates have a transient febrile reaction that may be caused by an increased serum concentration of proinflammatory cytokines. Aminobisphosphonates induce the production of certain proinflammatory cytokines in vitro, especially in cells of monocytic lineage. A unique feature of aminobisphosphonates is that they bind the Vgamma2Vdelta2 class of T cells, which are found only in primates, and stimulate cytokine production. The effects of aminobisphosphonates on other cells, including macrophages, are incompletely understood. We show in this study that treatment of murine macrophages with pamidronate, a second generation aminobisphosphonate, induces TNF-alpha production. Furthermore, pretreatment of murine macrophages with pamidronate before stimulation with IFN-gamma significantly augments IFN-gamma-dependent production of TNF-alpha. This pamidronate-mediated augmentation of TNF-alpha production results in sustained phosphorylation of the tyrosine residue at position 701 of STAT1 after IFN-gamma treatment. Our data suggest that this sustained phosphorylation results from inhibition of protein tyrosine phosphatase activity. We also show that pamidronate treatment increases TNF-alpha production in vivo in mice. Pamidronate-augmented TNF-alpha production by macrophages might be a useful strategy for cytokine-based anticancer therapy.

Implication of inflammation-related cytokines in the natural history of liver cirrhosis.

Increased serum concentrations of pro-inflammatory cytokines have been detected in patients with liver cirrhosis. However, their role in the natural history of cirrhosis and portal hypertension, in the absence of infection, and the prognostic significance of inflammation-related cytokines have not been reported. Our objective was the analysis of the prognostic value of inflammation-related cytokines in cirrhotic patients. Serum concentrations of tumor necrosis factor (TNF-alpha) and its soluble receptors I and II and interleukin 6 (IL-6), as well as mean blood pressure, plasma renin activity, aldosterone, vasopressin and norepinephrine concentrations were determined in 72 cirrhotic patients (Child-Pugh score: A 50%, B 33.3%, C 16.7%), without any evidence of infection, and in 25 healthy controls. Patients were followed up for a median of 35.9 (range 6-60) months. Increased concentrations of soluble TNF receptors were detected in cirrhotic patients when compared with healthy controls. TNF receptors and IL-6 concentrations were both significantly more elevated in advanced phases of cirrhosis (Child-Pugh score C vs B and vs A). Sixteen patients died as a related consequence of liver cirrhosis. Multivariant analysis demonstrated that Child-Pugh score, mean blood pressure and serum levels of TNF receptor I were associated with mortality. In addition to the classic factors implicated in mortality (Child-Pugh score and hemodynamic parameters), alterations in inflammation-related components are of prognostic significance in cirrhotic patients.

Effects of n-3 Fatty Acids on Serum Interleukin-6, Tumour Necrosis Factor-α and Soluble Tumour Necrosis Factor Receptor p55 in Active Rheumatoid Arthritis

We investigated the effects of a low n-6 fatty acid (FA) diet supplemented with fish oil on serum pro-inflammatory cytokine concentrations and clinical variables in patients with active rheumatoid arthritis (RA). Sixty patients were randomly assigned to receive a diet low in n-6 FAs and n-3 FAs supplement (fish oil group), a diet low in n-6 FAs and placebo (placebo group), or no special diet or intervention (control group). Serum cytokines and clinical and biochemical variables were evaluated at baseline and various timepoints. At week 18 the fish oil group had significant reductions in linoleic acid, C-reactive protein (CRP) and soluble tumour necrosis factor receptor p55 (sTNF-R p55), and significant elevations in eicosapentaenoic acid and docosahexaenoic acid compared with baseline. There were no significant differences in the clinical variables between the three groups. At week 24 there were significant reductions in interleukin-6 and TNF-alpha in the fish oil and placebo groups. Supplementation with n-3 FA and a low n-6 FA intake decreased serum sTNF-R p55 and CRP levels in patients with RA.

Periprocedural Kinetics in Serum Levels of Cytokines and Adhesion Molecules in Elective PTCA and Stent Implantation: Impact on Restenosis

To the Editor: Coronary restenosis after percutaneous transluminal coronary angioplasty (PTCA) remains a significant clinical problem occurring in ≈30% of patients even after wide accessibility and application of intraluminal stent implantation. Recent developments in stent design have made new techniques such as brachytherapy and drug-eluting stents available for clinical approaches; nonetheless, restenosis with its complex multifactorial genesis will continue to be an important problem with a major impact on long-term outcome of patients after coronary interventions. Several factors have been suggested to serve as predictors of the later occurrence of restenosis, but no serum parameter with predictive value has been clinically established so far. The pathophysiology of coronary restenosis has not been fully elucidated; however, an initial vascular injury is supposed to initiate a local inflammatory response that induces the recruitment of circulating immune-competent cells through an increased expression and translocation of several adhesion molecules such as P-selectin, E-selectin, and monocyte chemoattractant molecule-1 (MCP-1).1,2⇓ Migration of leukocytes, release of cytokines, and growth factors result in the induction of local inflammatory reactions and vascular remodeling.3 Patients with angina pectoris and acute myocardial infarction show a systemic inflammatory activation and elevated serum levels of vascular adhesion molecules.1,2⇓ Therefore, the local vascular inflammation seems to be accompanied by alterations of serum parameters exhibiting a systemic inflammatory response toward local tissue damage. We investigated periprocedural serum kinetics of cytokines (TNFα and IL-1β) and adhesion molecules (ICAM-1, VCAM-1, E-selectin and P-selectin) from baseline to 48 hours after elective PTCA and intracoronary stent implantation in patients with stable angina pectoris (CCS II-III). Forty patients with stable angina pectoris and a left ventricular ejection fraction of >45% admitted for elective PTCA and stent implantation were prospectively included in this study. Before and 48 hours after intervention, venous blood samples were drawn from …

Patients with non-immune chronic idiopathic neutropenia syndrome have increased splenic volume on ultrasonography.

Clinically detectable splenomegaly is rarely seen in patients with non-immune chronic idiopathic neutropenia syndrome (NI-CINS). Using ultrasound, we estimated splenic volume in 52 NI-CINS patients and 14 age- and sex-matched normal controls by determining the "corrected splenic index" (CSI) from the product of length, width and thickness of the organ expressed in cm3/m2 body surface area. We found that CSI was significantly higher in the group of patients compared to controls (202.8 +/- 82.0 vs. 133.8 +/- 28.1 cm3/m2, P=0.003), and that individual CSI values was inversely correlated with the number of circulating neutrophils (r=-0.5097, P < 0.0001). About 48.1% of the patients had CSI above 190 cm3/m2 body surface, representing the upper 95% confidence limit of values found in the controls. Patients also had increased serum concentrations of pro-inflammatory cytokines and chemokines mainly produced by activated macrophages (IL-1beta, TNF-alpha, RANTES and IL-8), as well as increased serum levels of soluble cell adhesion molecules derived from activated endothelium (sE-Selectin, sICAM and sVCAM). We hypothesize that the increased splenic volume in NI-CINS patients may be due to the accumulation of activated macrophages inside the spleen, possibly as the result of an unrecognized low-grade chronic inflammatory process. The nature of such an inflammation is unknown. A study was designed to search for viral or bacterial genomic material in patients' bone marrow stromal macrophages in which the unknown causal agent might be located.

Serum concentrations of proinflammatory cytokines in Graves' disease: effect of treatment, thyroid function, ophthalmopathy and cigarette smoking.

In the present study we have measured the concentrations of interleukin-6 (IL-6), soluble IL-6 receptor (sIL-6R), tumor necrosis factor-alpha (TNF-alpha), interleukin-1 beta (IL-1 beta) and IL-1 receptor antagonist (IL-1Ra) in the serum of patients with Graves' disease (GD). By multivariate analysis, we have evaluated the effect of antithyroid treatment, thyroid function, the presence or absence of active thyroid-associated ophthalmopathy (TAO), the patient's smoking habits and the relation to circulating anti-thyrotropin (TSH) receptor (TRAb) and anti-thyroperoxidase antibodies (TPOAb). We studied 84 GD patients, 51 untreated and 33 receiving methimazole (MMI) therapy. Twenty-three (45%) untreated patients and 18 (54%) patients on MMI had active TAO. We also studied 67 normal subjects as controls. Thirty-one GD patients (43%) and 16 controls (36%) were smokers. Serum IL-6 concentrations were significantly higher in both untreated patients (P<0.001) and treated patients (P<0.006), when compared with controls. Serum sIL-6R concentrations were significantly affected by treatment (P=0.001). Serum IL-1Ra concentrations were not different in GD patients, whether treated or untreated, compared with controls. Serum IL-6 concentrations were not influenced by thyroid function and there was a significant interaction between treatment and the presence of active TAO (P=0.003). In hyperthyroid patients with active TAO serum, sIL-6R concentrations were significantly higher than in those with inactive TAO (P=0.003). In untreated GD patients there was no significant effect of thyroid function and TAO activity on the serum concentrations of TNF-alpha and IL-1 beta. Serum IL-1Ra concentrations were not affected by the presence of TAO. Smoking had no effect on serum IL-6, sIL-6R, TNF-alpha, IL-1 beta and IL-1Ra concentrations, even in the presence of an active TAO. Serum concentrations of IL-6, sIL-6R, TNF-alpha and IL-1 beta and IL-1Ra were not different in patients with and without TRAb or TPOAb, in relation to either thyroid function, TAO activity or smoking. Our work shows that: (i) the proinflammatory cytokine pattern in GD is greatly influenced by antithyroid drug treatment; (ii) the increased circulating IL-6/sIL-6R concentrations observed in patients with active TAO may derive from the activation of humoral reactions in sites other than the thyroid; and, (iii) cigarette smoking has no effect on serum IL-1/IL-1Ra concentrations in TAO.

Differential immune responses to staphylococcal enterotoxin B mutations in a hydrophobic loop dominating the interface with major histocompatibility complex class II receptors.

Bacterial superantigens, such as staphylococcal enterotoxin B (SEB), can trigger acute pathologic effects in humans. A hydrophobic loop on the surface of SEB and other bacterial superantigens, centered around a conserved leucine (L45) residue, is essential for binding to class II major histocompatibility complex molecules. Single residue changes of wild type SEB, designated Q43P, F44P, or L45R, resulted in nonlethal proteins at a dose equivalent to 30 murine LD50 of SEB. Relative to SEB, the mutant proteins did not elevate serum concentrations of proinflammatory cytokines in mice and caused minimal proliferation of human lymphocytes. Anti-SEB titers of mice immunized with Q43P, F44P, L45R, or SEB were similar and protected 77%-100% of animals against a lethal SEB challenge. Levels of toxin-specific IgG1, IgG2a, IgG2b, and IgG3 in mice immunized with SEB, Q43P, or F44P were equivalent, but animals immunized with L45R had significantly elevated levels of IgG2a and IgG2b. Vaccines against staphylococcal superantigens should focus on this critical leucine residue.

Cytokines and Cardiomyocyte Death

Cytokines have been associated with the pathogenesis of acute coronary syndromes and chronic heart failure (CHF), which are both associated with cardiomyocyte loss. In CHF, increased serum concentrations of proinflammatory cytokines, including tumour necrosis factor α (TNF-α) and also soluble TNF receptor have been found. Both TNF and Fas-ligand have been able to induce programmed cell death (apoptosis) of cardio-myocytes in various experimental studies. In ischaemic conditions of the heart, increased serum levels of soluble Fas receptor have been found. The proinflammatory cytokines interleukin 1 (IL-1), IL-2 and interferon-γ can induce TNF production from target cells, including myocytes. TNF and some other cytokines are able to induce nitric oxide production, which depresses cardiac function and can induce apoptosis. However, anti-inflammatory cytokines such as IL-10, IL-4 and IL-13, secreted by T-helper type 2 lymphocytes and other cells, inhibit the production of proinflammatory cytokines. Preliminary studies suggest that cardiotrophin-1, produced by cardiomyocytes, is able to inhibit cytokine-induced cardiomyocyte apoptosis in vitro. As growth hormone is able to inhibit the production of proinflammatory cytokines in many cell types, it may also play an important role in the regulation of apoptosis induced by these cytokines. When the cytokine-induced pathways leading to altered gene expression of cardiomyocytes are understood, this knowledge may aid in the development of drugs that prevent progressive cardiomyocyte loss, in particular by inhibiting cytokine-induced apoptosis.

Precordial catch syndrome.

Seventeen cases of precordial "catch" are reported. The children varied in age from 8 to 16 years. The characteristics of this painful condition are discussed; in particular, the diagnostic indication of the site of the pain over an intercostal space, with the finger tip, is stressed. Anxiety in the child, or the parents, in relation to the complaint is discussed. Several methods of treatment are recommended.