Archives of Medical Research 32 (2001) 129–135

0188-4409/01 $–see front matter. Copyright © 2001 IMSS. Published by Elsevier Science Inc.PII S0188-4409(00)00272-1

ORIGINAL ARTICLE

Serum Immunoglobulins, IL-1

b

, IL-2, and IFN-

g

Gamma Level in Patients with Lymphedema Treated with Ortho-

b

-Hydroxy-Ethyl Rutosides (HR)

Diana G. Dlugovitzky de Schenquer* and Natalio Schenquer**

*Sección de Inmunología, Facultad de Ciencias Médicas, Universidad Nacional, Rosario, Argentina**Departamento de Bioquímica, Instituto de Especialidades, Rosario, Argentina

Received for publication September 1, 1999; accepted September 19, 2000 (99/144).

Background.

This investigation was undertaken to study the clinical characteristics andthe humoral immune pattern in lymphedema patients undergoing O-

b

-hydroxy-ethyl ruto-side therapy.

Methods.

A complete physical examination was performed on 27 lymphedema patients(postmastectomy and postphlebitis) and 17 healthy controls. Clinical evaluation of af-fected limbs was carried out concerning swelling, circumference, mobility, and tissue ten-sion, while immunologic studies consisted of serum IgG, IgA, IgM, C3, and 1-glycoprotein,interleukin-1 beta, interleukin-2, and IFN-

g

levels. Immunoglobulins and alpha-1-acidglycoprotein were assessed by radial immune diffusion (RID) and interleukin levels, byELISA (R & D Systems Kits, R & D Systems, Minneapolis, MN, USA). On admission,patients received O-

b

-hydroxy-ethyl rutosides (HR) 1 g/day during a period of 6 months.

Results.

These demonstrated several clinical, functional, and immunologic alterations inlymphedema patients. HR treatment produced a remarkable reduction in swelling, circum-ference, and tissue tension, and an improvement in mobility. In addition, edema was alsodiminished. The immunologic profile observed in untreated lymphedema patients is as fol-lows: IgG and

a

-1-acid glycoprotein concentration were slightly diminished in comparisonwith the control group, reaching normal levels after therapy. The difference among thesevalues was significant (

p

,

0.001). Contrariwise, normal C3 values were detected with asignificant posttreatment decrease (

p

,

0.001). Normal serum IgA and IgM levels in pa-tients with lymphedema before and after HR treatment are shown, and are similar to thoseof the control group. In addition, Il-1

b

and IL-2 values are higher in relation to control val-ues (

p

,

0.01), attaining normal concentration after therapy. Normal levels of IFN-

g

werefound in lymphedema patients, and there was no difference with control values. The con-centration of this cytokine was not influenced by HR. IL-1

b

and IL-2 concentrations werehigher than in controls (

p

,

0.001). Values reached normal levels after therapy (

p

,

0.01).

Conclusions.

Diverse clinical and immunologic alterations were evidenced in lymph-edema patients. The immunoglobulin profile and IL-1

b

and IL-2 levels were also modified.After HR treatment, their improvement was reflected by the favorable effect regarding thepreviously mentioned alterations. O-

b

-hydroxy-ethyl rutosides are effective both clini-cally and immunologically. © 2001 IMSS. Published by Elsevier Science Inc.

Key Words:

Lymphedema, Immunoglobulin, Cytokine, Treatment, Rutosides.

Introduction

Development of immunologic abnormalities in peripheraland dermal compartments of patients with lymphedema hasnot been fully elucidated. There is some evidence that tissuefluid stasis and inflammation may be involved in this regard(1,2). In fact, the inflammatory reaction appears to play animportant role, as inflammation can lead to changes usually

Address reprint requests to: Dra. Diana G. Dlugovitzky de Shenquer,Sección de Inmunología, Facultad de Ciencias Médicas, UniversidadNacional, Maipú 1340, Planta Baja, Depto. D, 2000, Rosario, Argentina.Tel.: (

1

54) (341) 448-4771; FAXES: (

1

54) (341) 480-4569 and 480-4558, ext. 249; or 449-4736; E-mail: schenquer@yahoo.com

130

Schenquer and Schenquer/ Archives of Medical Research 32 (2001) 129–135

reported in lymphedema patients, such as peripheral edema,increased concentrations of plasma proteins, and trapping offibroblasts and immunocompetent cells with their secretedproducts, as well as abundant collagen formation (3–7).

Leukocytes, in particular T lymphocytes and macro-phages, are sources of cytokines, and are influenced bythem, the latter responsible for diverse reactions that medi-ate inflammation and immunity (8). Thus, it follows thatabnormal cytokine synthesis may be a critical factor in theestablishment of inflammatory and immune disturbancesaccompanying vascular diseases (9,10).

With these considerations in mind, we decided to investi-gate whether the clinical degree of severity of lymphedemapatients (postmastectomy and postphlebitis) before and af-ter treatment is associated with a particular profile of immu-noglobulins, inflammatory proteins, and cytokine serumconcentrations. The investigation was developed by assess-ing the effect of treatment with ortho-beta-hydroxy-ethyl-rutosides (HR) on several clinical parameters, patient evolu-tion, and immunologic laboratory data.

Materials and Methods

Patients.

Seventeen untreated patients (aged 31–68 years,of both sexes) with lymphedema (some, of the legs (post-phlebitis) and the remainder of the arms (postmastectomy)due to breast cancer in women) were investigated. The samegroup of patients was studied after rutoside therapy.

Severity degrees of lymphedema patients were classifiedfrom I–V (4), and are characterized by a clinical and histo-logic status that refers to the severity and chronicity of thedisease, volume of the affected limb, and especially to thecellular subpopulations of the connective tissue and fibrosisthat increase according to the progression and severity oflymphostatic edema.

Patients of stages II and III of disease evolution weregrouped according to a classification cited by Pietravallo (4)that includes present clinical and histologic dysfunction re-garding degree of severity, chronicity, and volume of the af-fected limb and especially regarding connective cellularpopulations and fibrosis that increased with the progressionand gravity of the lymphatic edema.

Stage II.

Small, dilated lymphatic vessels were observed,more than in stage I in which dilation is slight. PMNs,monocytes, histiocytes, and necrobiotic cells, as well as cel-lular debris were increased. There is a collagen increase,both spread throughout and organized in the connective tis-sue. There are also fibroblasts around lymphatic vessels.

Stage III.

Connective and fibrotic bundles tightly surroundand distort the vessels. The connective tissue shows highlycellular areas attaining the subcutaneous tissue. Fibroblasts

are present. Dense collagen with distorted vessels is present,although there are regions of loose tissue.

Control group.

A normal control group, submitted to thesame studies, was also included.

Clinical studies.

In the first stage, the following severalclinical studies were performed to make a diagnosis and toascertain the etiology of the disease: 1) anamnesis, and 2)complete physical examination, focused on the affectedzones. Several aspects were considered in the affected legsand arms, as follows: 1. Swelling was measured by meansof water displacement (mL). Two measures of water vol-ume displaced by the affected limbs utilizing a water dis-placement plethysmograph device according to standard-ized procedures (11) were performed; 2. Circumference wasmeasured at the three points (precision: 0.1 cm) of the af-fected legs and arms. The circumference between lymph-edematous and the normal part of the limb was measuredwith a tape at the same site and at the same time; 3. In re-gard to mobility, the number of patients improving the mo-bility of the affected limb in comparison with the healthylimb function was evaluated, and 4. For tissue tension, therate of compression loss of the lymphedematous, and re-covery after rutoside treatment were assessed with a tono-meter. Evaluation was performed with a tonometer threetimes at each of two sites (distal and proximal) 10 cm fromthe elbow on the volar aspect of the arm or the dorsal aspectof the leg. Results are expressed in units of compression (ar-bitrary designation), and are the mean of three measure-ments. The higher the value, the softer the tissue (12,13).Taking into account that the normal limb was measured atthe same time as the affected limb, each patient was his/herown control. The compression units registered of the normaland affected limb were used to calculate the difference, pos-itive in case of the affected limb showing less compressionand the normal limb, or as negative with more compressionthan the normal limb.

The patient’s progress was assessed by measuring tissuetonicity in both the normal and the affected limb. Measureswere performed with the limb in horizontal position on a ta-ble, and the patient was told to allow the muscle to relax.

Values of the affected limb are expressed as percentagein comparison with the healthy limb to evaluate the influ-ence exerted by certain factors such as weather, activity, etc.Their corresponding values were determined before and af-ter treatment (14).

Edema characteristics that differed depending on theevolution of the disease were also established.

Paraclinical assays.

Paraclinical assays consisted of thefollowing: a) clinical laboratory studies; b) biological stud-ies to determine the presence of either infection or inflam-matory syndrome, and c) Stemer sign.

Schenquer and Schenquer / Archives of Medical Research 32 (2001) 129–135

131

Noninvasive methods.

These methods included 1) Doppler,2) Echodoppler, and 3) venous occlusion plethysmography.

Invasive methods.

These consisted of the following: 1)phlebography, and 2) lymphography.

Immunologic assays.

Serum samples were obtained 1 dayprior to HR treatment and after its completion (6 monthslater), and were stored at

2

70

8

C until used.IgG, IgA, IgM, C3, and

a

-1-acid glycoprotein (anotherimmune protein) serum concentration by immune radial dif-fusion (IRD) was assessed.

IL-1

b

, IL-2, and IFN-

g

serum levels by the so-calledELISA sandwich method were also tested. Commerciallyavailable enzyme-linked immunoadsorbent assay kits wereused for assaying these cytokines (Predicta ELISA forquantification of human IL-1

b

, interleukin-2, and inter-feron-

g

, Genzyme Corporation, Genzyme Diagnostics,Minneapolis, MN, USA). Briefly, serum samples were addedto the wells of microtiter plates precoated with anti-IL-1

b

,anti-IL-2, and anti-IFN-

g

mouse monoclonal antibodies tocapture the corresponding cytokine. After incubation at37

8

C for 2 h, the unbound components from the sampleswere removed by washing. Second polyclonal antibody (rat,rabbit) anti-human IL-1

b

, IL-2, and IFN-

g

were added andincubated for 1 h at 37

8

C. Subsequently, a third antibodygoat antirabbit IgG conjugated with peroxidase was addedand incubated for 1 h at 37

8

C. The reaction was developedafter adding the substrate and the stop solution that interruptcolor development. Finally, the color was measured with anELISA reader at 450 nm. Results were obtained by first cal-culating the average absorbance for each set of duplicatesamples and then analyzing the data by either manuallyplotting the standard curve or by entering the data into a pro-gram that generated a curve using linear regression analysis.

Control results.

The control data of protein and cytokinelevels were obtained from the same studies as conducted onhealthy volunteers. Serum control samples were obtainedand stored in the same manner as those of the patients.Quantification of immunoglobulin,

a

-1-acid glycoprotein,and cytokines was performed using the same method em-ployed with patient samples.

Treatment.

Patients received ortho(beta-hydroxy-ethyl ru-tosides) (Venoruton 1 g, Novartis Laboratory, Buenos Aires,Argentina) orally in the morning, 2 g/day during a period of6 months. Treatment efficiency was previously tested. Nev-ertheless, the drug tolerance of patients was verified toavoid any side effect or individual symptoms. The patientsdid not receive any other therapy that could act as an immu-nomodulator or modify the rutosides’ effect. This aspectwas strictly controlled. The variables were assessed in pa-tients both before and after treatment. Healthy volunteers(control group) were submitted to the same previously men-

Tab

le 1

.

Clin

ical

res

ults

of

lym

phed

ema

patie

nts

afte

r O

-bet

a-O

H-e

thyl

rut

osid

es tr

eatm

ent

Cir

cum

fere

nce

Thi

ghL

egSw

ellin

gT

issu

e te

nsio

nM

obili

ty

Lym

phed

ema

Pre-

HR

Post

-HR

Red

ucPr

e-H

RPo

st-H

RR

educ

Pre-

HR

Post

-HR

Red

ucPr

e-H

RPo

st-H

RR

educ

Pre-

HR

Post

-HR

Red

uc

Post

-phl

ebiti

s58

.8

6

6.9

53.6

6

6.1

91.5

6

10.

536

.9

6

4.6

34.2

6

4.8

91.2

6

16.

927

6

2.6

22

6

5.2

82

6

15.

728

6

7.2

18

6

9.8

65.5

6

17.

927

6

11

31

6

14

90.5

6

26

Arm

Fore

arm

Pre-

HR

Post

-HR

Red

ucPr

e-H

RPo

st-H

RR

educ

Pre-

HR

Post

-HR

Red

ucPr

e-H

RPo

st-H

RIn

cr.

Pre-

HR

Post

-HR

Rec

up

Post

-mas

tect

omy

30

6

15.

419

.8

6

8.9

60.8

6

22.

731

.8

6

9.3

25.8

6

9.3

81.1

6

9.4

33

6

4.1

24

6

3.1

72

6

8.9

32

6

10.

323

6

17.

368

6

16.

429

6

10.

733

6

16.

687

.2

6

17.

6

The

exp

ress

ed v

alue

s ar

e m

ean

6

sta

ndar

d de

viat

ion

(mea

n

6

SD

); d

iffe

renc

es b

etw

een

pre-

HR

and

pos

t-H

R w

ere

eval

uate

d by

Stu

dent

t

test

; Pre

HR

; Pre

trea

tmen

t with

HR

; Pos

t-H

R: P

osttr

eatm

ent w

ithH

R; M

obili

ty: R

ecov

ery

afte

r tr

eatm

ent,

6

SD o

f 15

; Red

uc: R

educ

tion

of c

ircu

mfe

renc

e, s

wel

ling,

or

tissu

e te

nsio

n; C

ircu

mfe

renc

e: A

sses

smen

t of

thre

e an

atom

ic p

oint

s of

leg

or a

rm; R

ecup

: Rec

uper

atio

n.

132

Schenquer and Schenquer/ Archives of Medical Research 32 (2001) 129–135

Immunologic data.

Several alterations were observed re-garding Igs, C3, and

a

-1-acid glycoprotein serum levels.These values are shown in Table 2. IgG and

a

-1-acid glyco-protein concentration was slightly diminished previous totreatment in comparison with the control group, reachingnormal levels after therapy. The difference between thesevalues was significant (

p

,

0.001). To the contrary, normalC3 values were detected before HR treatment, with a signif-icant post-treatment decrease (

p

,

0.01).Table 2 displays normal serum IgA and IgM levels in

lymphedema patients, which are modified when comparedto those of the control group values or after HR treatment.IL-1

b

, IL-2, and IFN-

g

concentrations are presented in Ta-bles 4 and 5. Il-1

b

and IL-2 yielded higher concentrations inrelation to control values (

p

,

0.01) and to posttreatment es-timations; they attained normal values (

p

,

0.01) after ther-apy. Normal levels of IFN-

g

were found in lymphedema pa-tients; there was no difference with control values. Theconcentration of this cytokine was not influenced by HRtreatment. Severity of lymphedema is classified in fourstages according to swelling intensity, edema, and in partic-ular by histologic alterations such as connective tissue den-sity and number of collagen fibers. Intensity of these char-acteristics increases from stages I–IV. The patients withlymphedema in the present study belonged to stages II orIII. Serum immunoglobulins (Table 3) and cytokines (Table5) of patients with lymphedema classified as stages III or II,the most prevalent conditions before and after treatment, re-spectively, exhibited a difference in these parameters. Thus,IgG and alpha-1-acid glycoprotein data were significantlydecreased in lymphedema stage III in comparison with con-trols (

p

,

0.01 and

p

,

0.001, respectively). C3 was signifi-cantly higher in stage II than in stage III (

p

,

0.001). Therewere no significant differences in IgM values betweenstages II and III, which did not differ from controls. In addi-

tioned immunologic assays. Both patients and healthy vol-unteers signed an informed consent. This investigation con-forms with the principles outlined in the HelsinkiDeclaration (15).

Statistical analysis.

The results of immunoglobulin and

a

-1-acid glycoprotein and differences in cytokine valueswere analyzed by paired Student

t

test. The 0.05 level ofsignificance was accepted.

Results

The results demonstrated several clinical, functional, andimmunologic alterations in patients with lymphedema as re-flected in the evaluated parameters.

The disease is characterized by peripheral edema withaccumulation of metabolic products, trapped circulatinglymphocytes, and macrophages with production of collagenand fibrosis.

Some changes were also observed in these estimationsafter HR treatment.

Clinical results.

Data revealed an important swelling, cir-cumference and tissue tension increment, and decrease ofmobility in untreated patients with lymphedema.

Clinically, the patients had a favorable evolution with animportant edema decrease; chronic venous insufficiencysymptoms were also diminished. The results of the evalu-ated clinical parameters are shown in Table 1. Data con-cerning diminished swelling, circumference reduction, in-creased mobility, and tissue tension reflect a favorableclinical evolution. There was good tolerance of HR therapyand no unfavorable reactions.

Table 2. Serum levels of IgG, IgA, IgM, C3, and a-1 acidglycoprotein in lymphedema patients treated with O-beta-OH-ethyl rutoside (HR)

Serum levels IgG IgA IgM C3a-1-acid

glycoprotein

Lymphedema pretreatment (n 5 27) 659 6 0.5 260.64 6 28.56 145.76 6 46.9 124.58 6 16.97 56.76 6 20.64Lymphedema posttreatment (n 5 27) 1000.7 6 82.44a 259.82 6 25.85 147.76 6 45.43 98.70 615.16a 82.65 6 21.02b

Difference 341.7 0.82 2 25.88 5.59Normal controls (n 5 18) 1067.25 6 97.39b 211.75 6 72.17 126.5 6 55.99 133.87 6 31.8 90.25 6 24.31b

The values are mean 6 standard deviation (mean 6 SD); ap ,0.001; bp ,0.01.

Table 3. IgG, IgA, IgM, C3, and a-1-acid glycoprotein serum levels in lymphedema patients of different degrees of severity

Serum levels IgG IgA IgM C3a-1-acid

glycoprotein

PatientsDegree II (n 5 12) 623 6 0.8a 260 6 21.2 136 6 46.1 121.7 6 16.1 68.35 6 20.04c

Degree III (n 5 15) 684 6 0.52a 270 6 19.7 131.0 6 18.3 80.3 6 13.8a 50.6 6 21.07c

Normal controls 1067.25 6 97.39b 231.785 6 72.17 126.5 6 55.99 133.87 6 31.08b 90.25 6 24.31c

The values are mean 6 standard deviation (mean 6 SD); Student t test; ap ,0.01; bp ,0.001; cp ,0.05

Schenquer and Schenquer / Archives of Medical Research 32 (2001) 129–135 133

tion, differences in cytokine values as related to disease se-verity (Table 5) were observed, mainly IL-1b and IL-2.Both parameters are higher in stage III than in stage II(p ,0.01).

Considering IL concentrations in regard to disease sever-ity, it was observed that patients at stage III presentedhigher IL-1b and IL-2 values in comparison with controls(p ,0.001), and that these differences decreased in stage II.IFN levels were normal in both stages III and II (Table 5).

Discussion

This study showed diverse clinical, functional, and immu-nologic alterations in patients with lymphedema, as well asmodifications of these parameters regarding patient im-provement.

Edema, swelling of the affected arm or leg, diminishedmobility, and decreased tissue tension were deemed impor-tant by the evidence. In young people, the extremity devel-oped without modifications of skin texture, but in older sub-jects, fibrotic skin changes were observed. These phenomenaare probably related to a subclinical inflammation of theskin. We have recently reported the histologic changes de-tected with immunohistochemical techniques, characterizedby proliferation of keratinocytes and fibroblasts and accu-mulation of lymphocytes and macrophages (6) that may bethe result of the production of cytokines such as IL-1b. Sim-ilar observations were also described by Olszewski et al.,who have investigated these subacute inflammatory changesof the skin in patients with filarial lymphedema of the legs(16,17). Therapy with HR not only reduced the swelling,circumference, and tissue tension of the affected extremity,but also improved mobility.

There are previous studies in the literature that demon-strate that flavonoids, alone or with compressive tech-niques, are a useful tool in lymphedema therapy. A clinicalsymptomatologic improvement was verified in several re-ports (18,19), as well as a remarkable diminution of swell-ing and volume of the affected limb. In addition, their anti-inflammatory, antioxidant, and antiaggregating action,which could be due to leukotriene and prostaglandin synthe-sis inhibition, oxygen-reactive intermediates, and nitric ox-ide has been studied (20–22).

Flavonoids have shown a lower release of fluid to the in-terstitium, in addition to a decrease in protein deposits andlower lymphocyte activity. It has been proposed that the di-versity of these compound activities could be linked to aninteraction with endothelin; the latter, however, has not yetbeen demonstrated (23).

In the present study, diminished IgG and a-1-acid glyco-protein concentration was detected in untreated patients.This finding could be explained by stagnant lymph inlymphedema patients, which has a high protein concentra-tion (24), or by changes in lymph protein concentrations ofdifferent molecular weights (16,25).

HR treatment induced normal levels of these immuneproteins. Several reports have described the beneficial effectof benzopyrones in patients with lymphedema and high pro-tein levels (25,26). C3 concentration that was normal beforetherapy was decreased after same. Complementary C3 lev-els gradually decreased over a 1-month period of HR treat-ment, reaching statistical significance compared with con-trols. This observation suggests that in lymphedema, a highrate of C3 synthesis is induced and that this process does notcontinue during the posttreatment phase.

IgA and IgM concentration was normal and these valueswere not influenced by the therapy.

Table 4. Levels of IL-1b, IL-2, and IFN-g in patients with lymphedema

Cytokines pg/mL in lymphedemapatients

IL-1b IL-2 IFN-g

Pre-HR Post-HR Pre-HR Post-HR Pre-HR Post-HR

Postphlebitis (n 5 13) 236 6 18.1 109.8 6 6.9a 172.8 6 8.6 170 6 6.9a 170 6 15 158.1 6 13Postmastectomy (n 5 14) 216 6 16.7 101.7 6 7.8 166.2 6 9.7 69.6 6 7.6a 153 6 21 162.3 6 14Normal controls (n 5 18) 91.7 6 4.7b 58 6 2.7b 160 6 19

The values are mean 6 standard deviation (mean 6 SD); Pre-HR: pretreatment; Post-HR: posttreatment; ap ,0.01; bp ,0.001.

Table 5. Cytokine serum levels in lymphedema patients with different degrees of severity

Cytokine (pg/mL)lymphedema

IL-1b IL-2 IFN-g

Degree II Degree III Degree II Degree III Degree II Degree III

Postphlebitis (n 5 13) 167.3 6 10.1 278.1a 6 10.2 96.4 6 7.2 178a 6 11.3 168 615.4 156 6 12.5Postphlebitis (n 5 14) 179.4 6 8.4 246a 6 15.3 76.4 6 6.9 102a 6 8.7 166 617.2 153 6 13.5Postphlebitis (n 5 18) 91.4 6 4.7b 58b 6 2.7 160 6 19

Student t test; ap ,0.01; bp ,0.001.

134 Schenquer and Schenquer/ Archives of Medical Research 32 (2001) 129–135

The development of lymphedema caused by filaria,which affects about 90 million people throughout the world,led several authors to investigate the specific and unspecificimmune response in patients with filaria (1,5,9,10,18,20,21,27).It has been demonstrated that the development and progressof filarial disease is a consequence of several immune reac-tions; nevertheless, it is still unclear which phenomena areproduced by the inflammatory and fibrotic processes char-acteristic of the lymphedema, and which are a consequenceof the immune response to the parasite.

Nonfilarial lymphedema, which is prevalent in SouthAmerica, was also investigated in depth. Results support thehypothesis that development of an obstructive disease of thelymph glands possesses an immune component in these pa-tients (7). Other studies demonstrated the relationship ofT- and B-cell responses in active infection, clinical manifes-tations in filarial and nonfilarial lymphedema, and specificand nonspecific lymphocyte proliferation, which was higherin elephantiasis patients as well as in amicrofilaremics (9).These findings provide evidence for clinical and laboratorymanifestations of this disease being caused by different im-mune responses.

As already described (6), tissues of the affected extremityare infiltrated with lymphocytes and macrophages probablyrecruited from the peripheral compartment. This concentra-tion of immune cells may relate to the high level of IL-1b inthe peripheral compartment. Because adequate function ofmacrophages and T-lymphocytes is essential for both pro-tective immunity and inflammatory reactions in response tofactors causing lymphedema, it is reasonable to assume thatthis clinical outcome is largely dependent on the type of im-mune response that develops in this disease. The altered pat-tern of immune proteins and high activity IL-1b suggestspersistence of an ongoing local inflammatory process.

Increase of IL-2 is responsible for T-cells differentiation,and may act to recruit other T-cells subsets, gd T-cells, andCD8 T-cells that probably exert an additional role in thegenerated immune response, although IFN-g levels werenot modified as either a consequence of lymphedema or HRtreatment. T-lymphocyte and macrophage activation that isproduced in tissue lesions induces a variety of changes suchas collagen deposits and fibrosis.

Beyond undesirable side effects, cytokines might play aregulatory effect to reduce skin and tissue alterations (28).Factors determining this profile of circulating cytokines arestill poorly understood, while the presence of locally operatinginfluences, such as those delivered by stimulated macro-phages that seem to play an important role, may account forthis process.

Differences in the pattern of circulating immunoglobu-lins, a-1-acid glycoprotein, C3, and cytokines reported herecan help to explain data registered in our and other laborato-ries (25,29,30) in which postphlebitis and postmastectomylymphedema is accompanied by diverse alterations in hu-moral immune proteins and interleukins.

AcknowledgmentsFinancial support was provided by the Novartis Laboratory, Bue-nos Aires, Argentina.

References1. Mak JW. Advances in immunology and immunopathology of lymphatic

filariasis. SoutheastAsian J Trop Med Public Health 1993;24(Suppl 2):76.2. Jiménez Cossio JA. Diagnóstico y tratamiento de los linfedemas.

Madrid, Spain: Lab. Uriach;1987.3. Foldi E, Foldi M, Clodivs L. Das Lymphödemchaos. Phlebol Provtol

1987;16:89.4. Pietravallo A. Linfedema: tratamiento farmacológico, fisioterapeútico

y quirúrgico. Buenos Aires, Argentina: Lab Phoenix;1993. p. 22.5. Lal RB, Dhawan RR, Ramsy RM. C-reactive protein in patients with

lymphatic filariasis. J Clin Immunol 1991;11:46.6. Dlugovitzky D, Schenquer N, Curto O. Phlebologie. Altérations des

niveaux des cellules immunocompetentes et des modifications histi-ologiques chez des patients avec du limphoèdeme traités avec l’orto-beta-hydroxi-ethylrutoside (HR). Phlébologie 1998;51(1):57.

7. Lammie PJ, Addiss DG, Leonard G, Hightower AW, Eberhard ML.Heterogeneity in filarial specific immune responsiveness among pa-tients with lymphatic obstruction. J Infect Dis 1993;167(May)(5):1178.

8. Libby P, Clinton SK. Cytokines as mediators of vascular pathology.Nov Rev Fr Hematol 1992;34:547.

9. Yazdanbakhsh M, Paxton WA, Kruize YC, Sartono E, Kurhiawan A,Van-het-Wout A, Slekirt ME, Partono F, Maizels RM. T-cell respon-siveness correlates differentially with antibody isotype levels in clini-cal and asymptomatic filariasis. J Infect Dis 1993;167(4)(Apr):925.

10. King CH, Ottesen EA, Nutman TB. Cytokine regulation of antigendriven immunoglobulin production in filarial parasite infection in hu-mans. J Clin Immunol 1990;85:1810.

11. Swedborg I. Volumetric estimation of degree of lymphoedema and itstherapy by pneumatic compression. Scand J Rehab Med 1997;9:131.

12. Clodius L, Deak L, Piltier NB. A new instrument for the evaluation oftissue tonicity in lymphedema. Lymphology 1976;9:1.

13. Piller NB, Clodius L. The use of a tissue Tonometer as a diagnostic aidin extremity lymphoedema: a determination of its conservative treat-ment with benzopyrones. Lymphology 1976;9:127.

14. Piller NB, Morgan RG, Casley Smith JR. A double blind cross over trialof O-(beta-hydroxyethyl)-rutosides (benzo pyrones) in the treatment oflymphoedema of the arms and legs. Br J Plast Surg 1988;41:20.

15. Declaration H. Br Med J 1964;11:177.16. Olszewski WL. Peripheral lymph: formation and immune function.

Boca Raton, FL, USA: CRC Press;1985.17. Olszewski WL, Janal S, Manukaran G, Lukomska B, Kubieka V. Skin

changes in filarial and non filarial lymphedema of the lower extremi-ties. Trop Med Parasitol 1993;44(1):40.

18. Pecking AP. Evaluation by lymphoscintigraphy of the effect of mi-cronized flavonoid fraction (Daflon 500 mg) in the treatment of upperlimb lymphedema. Int Angiol 1995;14(Sep 3)(Suppl 1):39.

19. Neumann HA, Van Den Broek MJ. A comparative clinical trial ofgraduated compression stockings and O-(beta-hydroxyethyl)-rutosides(HR) in the treatment of patients with chronic venous insufficiency. ZLymphol 1995;19(Aug)(1):8.

20. Abad MJ, Bermejo P, Villar A. The activity of flavonoids extractedfrom Tanacetum microphyllum DC. (Compositae) on soybean lipoxyge-nase and prostaglandin synthetase. Gen Pharmacol 1995;26(Jul)(4):815.

21. Robak J, Gryglewsky RJ. Bioactivity of flavonoids. Pol J Pharmacol1996;48(6):555.

22. Lee RW, Macintyre JP, Taylor GR, Wang E, Plante RK, Tam SS, PopeBL, Lau CY. Tepolaxin enhances the activity of an antioxidant, pyrrolidinedithiocarbamate, in attenuating tumor necrosis factor alpha-induced apop-tosis in WEHI 164 cells. J Pharmacol Exp Ther 1999; 289(Jun)(3):1465.

Schenquer and Schenquer / Archives of Medical Research 32 (2001) 129–135 135

23. El Internista. New York: McGraw-Hill;1997. p. 338.24. Olszewski WL, Engeset A. Lymph flow and composition in normal

conditions and in patients with lymphedema. In: Nishi MS, Uchino S,Yabuki R, editors. Progress in lymphology. Proc. XII Int. Congress ofLymphology. Tokyo: Elsevier;1989.

25. Olszewski WL, Jamal S, Lukomska B, Manokaran G, Grzelak I. Im-mune proteins in peripheral tissue fluid-lymph in patients with filariallymphedema of the lower limbs. Lymphology 1992;25:166.

26. Casley-Smith JR, Casley-Smith Judith R. High proteins oedemas andbenzo-pyrones. Baltimore, MD, USA: Lippincott Sidney; 1986.

27. Estambale BB, Simonsen PE, Vennervald BJ, Knight R. Bancroftian

filariasis in Kwale, District of Kenya. Humoral immune responses tofilarial antigens in selected individuals from an endemic community.Ann Trop Med Parasitol 1994;88(Apr)(2):153.

28. Pober JS. Cytokine-mediated activation of vascular endothelium:physiology and pathology. Am J Pathol 1988;133:426.

29. Freedman DO, Nutman THB, Jamal S, Kumaraswami V, Ottensen EA.Selective regulation of endothelial cell class 1 MHC expression bycytokines from patients with lymphatic filariasis. J Immunol 1989;142:653.

30. Olszewski WL, Engeset A. Immune proteins, enzymes and electrolytesin peripheral lymph. Lymphology 1978;11:156.