0022·202X/ 82/7902·0104$02.00/ 0 TH E JOURNAL OF INVESTIOATIVE DERMATOLOGY , 79:104-108, 1982 Copyright © 1982 by The Williams & Wilkins Co.

Vol. 79, No.2 Printed in U.S.A.

On the Mechanisms of Enhanced Monocyte and Neutrophil Cytotoxicity in Severe Psoriasis

TROELS RERLlN, M.D., NIELS BORREGAARD, M.D., AND KNUD KRAGBALLE, M.D.

Department of Dermatology, Marselisborg Hospital, (TH & KK), and Department of Medicine and Haematology, Aarhus Amtssygehus (NB), Aarhus University, Aarhus, Denmark

Monocyte and neutrophil function assessed as anti­body-dependent cell-mediated cytotoxicity (ADCC) us­ing IgG-sensitizing human erythrocytes as target cells was enhanced in patients with severe psoriasis when compared to .healthy controls. We found significant cor­relation between increased monocyte ADCC and in­creased neutrophil ADCC. No differences in basal cAMP levels and cAMP responses during initiation of the ADCC reaction was observed between psoriatics and normals. Also degranulation determined as lysozyme release dur­ing ADCC was normal. In contrast, the increase in ADCC was significantly correlated to an enhanced hexose monophosphate shunt activation in the effector cells during the cytotoxic reaction. Activity of enzymes re­sponsible for the respiratory burst was not altered in psoriasis since superoxide production after stimulation with phorbol myristate acetate was normal. Likewise, oxygen consumption and degranulation following phag­ocytosis of opsonized zymosan particles in neutrophils was found normal in psoriasis. Since monocytes showed increased binding of IgG-sensitized erythrocytes these data indicate that the enhanced monocyte and neutro­phil ADCC is caused by an enhancement of the respira­tory burst possi1;>ly induced by increased Fc receptor activity.

Increased attention has recently been focused on the en­hanced in vitro functions of monocytes and neutrophils from patients with psoriasis. Thus, enhanced chemotactic, phago­cytic, and cytotoxic activities have been found in phagocytes from psoriatic subjects [1-6). Since agents elevating cAMP levels depress chemotaxis and phagocytosis [7,8], and since cGMP has the opposite effect [9], it has been suggested [1-5], although not experimentally confIrmed, that the cause of the enhanced leukocyte function might be a decreased cAMP / cGMP ratio. In a previous study we compared the monocyte and neutrophil in vitro functions determined as antibody-de­pendent cell-mediated cytotoxicity (ADCC) to the intracellular content of cyclic nucleotides [6], and we demonstrated that the enhanced ADCC found was neither explained by an abnormal cyclic nucleotide metabolism nor by the presence of stimulating serum factors. This study investigates the events of the cyto­toxic process: initial cAMP increments during ADCC [10],

Manuscript received September 30, 1981; accepted for publication January 18, 1982.

This work was supported by the Danish Medical Research Council (grant no. 12-1784 + 12-0334).

Reprint requests to: Troels Herlin, M.D., Department of Dermatol­ogy, Marselisborg Hospital, DK-8000 Aarhus.c, Denmark.

Abbreviations: ADCC: antibody-dependent cell-mediated cytotoxicity cAMP: cyclic adenosine 3'-5' -monophosphate FCS: fetal calf serum HMPS: hexose monophosphate shunt NBT: nitroblue tetrazolium PMA: phorbol myristate acetate R: residue STZ: serum treated zymosan

lysosomal enzyme release and activation of the hexose mono­phosphate shunt [11,12]_

MATERIALS AND METHODS Subjects

Twelve patients (7 males and 5 females) with psoriasis vulgaris were investigated. The age range was 24 to 58 yr with a median of 32 yr. The patients had not received systemic medication for their psoriasis within 1 yr before the investigation and were without cW'rent infections. T he patients were treated with topical steroids and/or tru-. All had severe psoriasis with more than 25% of the skin sW'face affected. The experi­ments were performed with one patient and one healthy control inves­tigated in parallel. Controls were 12 healthy staff members (6 males and 6 females) aged from 23 to 49 yr with a median of 33 yr.

Isolation of Monocytes and Neutrophil Leukocytes

Monocytes and neutrophils were isolated from heparinized venous blood as described elsewhere [11,13]. The purity of the monocyte suspension as judged by nonspecific esterase staining was 93% ranging fTom 89% to 97%_ Occasionally, 1 to 3% granulocytes were present, the rest were lymphocytes. Judged by morphological criteria the isolated neutrophils were more than 97% pure.

Antibody·dependent Cell· mediated Cytotoxicity

The cytotoxicity assay was performed with minor modifications of the procedure previously described [14]. The tests were set up in duplicates and all cells were resuspended in Hanks balanced salt solution (HBSS) with 10% medium RPMI 1640 (Gibco, Grand Island, N .Y.), 5% heat-inactivated fetal calf serum (FCS), 100 lU/ ml penicillin and 100 Jlg/ml streptomycin, and 25 mM Hepes. Different tru-get cells were used in monocyte and neutrophil ADCC. For the monocyte ADCC human rhesus D-positive erythrocytes were used as tru-get cells. These erythrocytes (50 JlI of 1000 x 10"/ml) were labelled with sodium-51 chromate and sensitized with 150 III of 100% hyperirnmune anti -D antiserum. The monocytes (100 JlI of 1 x 10"/ml) were mixed with pretreated erythrocytes (200 l.tI of 6 x 10" /ml) . As target cells for the neutrophil ADCC assay o'Cr-labeled human type A erythrocytes were employed. The neutrophils (100 III of 1 x 10"/m!) were incubated with the type A erythrocytes (100 JlI of 6 x lO"/m!) and 100 III of 40% hyperirnmune anti-A antiserum.

104

The ADCC reaction was initiated after centrifugation of the mixtW'e at 150 g for 1 min, the cells were incubated at 37°C for 30 min and finally half of the supernatan~ was withdrawn. This supernate, C, was counted together with the residue (R) in a gammacounter (Mini Gamma, LKB). The chromium release was determined as follows:

% 5'Cr-release C cpm x 2 x 100

C cpm + Rcpm

For each reaction the specific c, 'Cr-release was obtained by subtract­ing the release in control tubes fTom the release in tubes containing phagocytes. The results were expressed as the number of target cells lysed per phagocytes:

number of targets X % specific r"Cr-release

number of phagocytes x 100

Hexose Monophosphate Shunt (HMPS) .

The respiratory burst during the ADCC reaction is reflected by the activation ofHMPS that can be assessed by the conversion of glucose­l-"C to '·C02 [11]. T he Embden-Meyerhof pathway is unchanged during ADCC since no increase in '·C02 from glucose-6-'·C is ob­served [11]. The cells were incubated in the presence of 1.2 mM (1-'·C)-glucose (specific activity 2.7 mCi per mmole). After incuba~ioll

Aug. 1982

for 30 min the reaction was terminated by injec ting 100 ,.u 1 N NaOH through the caps in to the cell mixtures. The tube contents were re moved after 1 hI' and placed in to siliconized conical flas ks conta ining 800 }.tI 1 N HCI and immediately hereafter t he fl asks were sealed with a rubber cap. After mixing with H CI, the dissolved CO2 was driven out and a bsorbed by 400 }.tI 1 N NaOH placed in a centre well in the bottom of the fl asks. After standing overnight on a sha king bath the content of the well representing the '''C02 activity was withdrawn, dilu ted in 1.5 ml H,O a nd counted in 10 ml Ready-Solv (Beckman). The ','CO, ac tivity produced by unstimu lated cells was subtracted from the values obtained with stimulated cells.

Degranulation

Assay mixtures were incubated as described for the ADCC assay. After 30 min at 37° 200 ~tl of the supernatant was wi thdrawn for determining of lysozyme. The residue was ul trasonica lly disrup ted in 0.2% Triton X-IOO (Sigma, St. Louis). Lysozyme (Tom both superna­tant a nd cell pellet was measured turb idometrically by the lysis of micrococcus lysodeikticus [15). Lysozyme release was exp ressed as the percentage of tota l lysozyme content and the sponta neous release from unstimulated cells was sub tracted to obtain specific lysozyme release. Beta-glucuronidase from azurophil granules is liberated dUTing ADCC (Herlin et ai , to be published) , however, it is not demonstrable during incubation with RPM! 1640 used here since phenolph thalein in terferes with the determination of the enzyme.

Degranulation during complement-mediated neutrophil phagocyto­sis was performed by addit ion of 0.5 mg/ml of serum-treated zymosan particles (S igma) to neutrophils (3 x lO"/ ml) resuspended in HBSS wi th 25 mM Hepes and 5 mM glucose. Phagocytosis was arrested in ice­cold 1 mM iodoacetamide. The tubes were centrifuged and supernata nts removed for enzyme determination. The cell pellet was ul trasonically disrupted in 0.2% Triton-X-lOO. Lysozyme was measured as described above a nd betaglucuronidase was dete rmined afte r incubation with phenolphthalein betaglucm onic acid for 18 h at 37°C [16).

Cyclic AMP L evels during ADCC

Monocytes and neutrophils were I'esuspended in HBSS with 2.5% FCS and adjusted to 15 x 10';/ ml. The same target cells were employed as for the ADCC assay. Addi t ion of sensit ized erythrocytes was added to either monocytes or neut rophils as previously described [10). At the time indicated the reaction was stojJped and cAMP was extracted with addition of 60 III of boiling potassium phosphate buffer as previously descri bed [17].

Cyclic AMP was determined by a competitive protein binding method using a cAMP assay kit (Radiochemica l Centre, Amersham, England) .

Erythrocyte- antibody Rosetting T est

R hesus D-positive erythrocytes (50 ~ll of 1000 x 10"/ ml) were eith er sensitized with 150 III of 100% a nt i-D antiserum as described for ADCC or with 150 III of either 25%, 12.5% or 6.25% antiserum. To 100 III of monocytes (1 x lO';/ ml) 200 III of sensitized ery throcytes (20 X lO"/ ml) were added and the cell mixture was cent rifuged (150 g for 3 min) and incubated for 5 min at 37°C. The reaction was stopped by plac ing the tubes in an ice-water bath . Acridine orange was added a nd under tluorescent-microscope monocyte ' wi th at least 3 erythrocytes a ttached were coun ted as rosettes.

0 , consumption during Pha.gocytosis

Neutrophils (1 ml of 3 X lO"/ ml) were stimulated by addi t ion of 0.5 mg/ ml of zymosan t rea ted with fresh serum prepared as described in [18]. O2 consumption was measured in a rapidly st irred thennostat ized chamber (37°C) with a Clarke- type oxygen electrode bu il t. under the stirrer (Rank Bros, Bottisham, E ngland) attaching the electrode to a Beckman recorder.

Superoxide Production.

To monocytes and neutrophils (600 ~ll of 1 X lOn/ ml) 5.0 Ilg/ ml phorbol myristate acetate (PMA) was added a nd superoxide generation was measured as previously described [19] by followin g the increase in absorbance at 550 nm in the presence of 0.2 mM cytochrome c.

Statistical Analysis

For statistical evaluation a Mann-Whitney test for 2 a mples was employed considering a p -value below 0.05 as significant. Spearman's coefficient of rank was applied in the correlation studies.

CYTOTOXICITY IN SEVERE PSORIASIS 105

RESULTS

Monocytes a nd neutrophils isola ted from patients with severe psoriasis vulgaris exhibited enhanced in vitro function deter­mined by their antibody-dependent cell-mediated cytotoxicity. In psoriasis the mean monocyte ADCC was 137% (p < 0.02%) and neutrophil ADCC was 154% (p < 0.01) of control values (Table I). Nine ofthe 12 patients had ADCC values above 120% of controls investigated on the same day. Increased monocyte ADCC parallelled increased neutrophil ADCC (R = 0.7483, P < 0.02, n = 12).

We then tried to delineate the processes involved during ADCC. From Table I it is seen that activation of h exose monophosphate shunt dming monocyte and neu trophil ADCC was significantly increased in psoriatics (p < 0.01 and p < 0.05, respective ly) whereas HMPS activity in unstimulated cells was not different from normal (954 ± 162 cpm versus 910 ± 184 cpm for monocytes and 904 ± 181 cpm versus 922 ± 175 cpm for neutrophils). Furthermore, increased HMPS activation was significantly related to increased ADCC by both monocytes (Fig 1, I' = 0.8392, p < 0.01) and neutrophils (Fig 2, r = 0.7063, P < 0.02) . Dming ADCC degranulation of leukocytes from psoriatics measured as lysozyme release did not differ signifi­cantly from normals (Table I). We found no r elation between degranulation and ADCC in psoriasis (r = 0.1235, N.S., for monocytes and r = 0.2036, N.S. , for neutrophils). Total lyso­zym e content in monocytes and neutrophils from psoriatic subjects was normal.

Cyclic AMP levels were recorded during the initial phase of ADCC in order to see if a possible impairment of cyclic neu­cleotide metabolism in psoriasis migh t be expla natory for th e enhanced ADCC observed. Because of shortage of cells this investigation was not performed in 2 of the patients studied. From Fig 3 and Fig 4 it is seen that normal cAMP values are present in unstimulated monocytes and neutrophils from the psoriatics. Addition of sensitized rhesus D-positive erythrocytes to the monocytes caused a rapid a nd transient increase in cAMP levels (Fig 3). The maximal increase was the same in

TABLE I. Events during antibody-dependent cytotoxicity mediated by m.onocytes a.nd neutrophils in. severe psoriasis

Psori asis Healthy co ntrols

Monocytes Cytolysis (erythro- 1.56 ± 0.14 ** 1.14 ± 0.11

cytes lysed per monocyte)

HMPS (cpm 14C02 2455 ± 294 " 1389 ± 142 from (J -"'C)-glu -cose)

Lysozyme release 19.8 ± 1.5 16.3 ± 1.8 (%)

Maximal increase in 14.2 ± 2.9 11.5 ± 2.4 cAMP (pmol per 10' cells)

Neutrophils Cytolysis (erythro- 1.14 ± 0.08" 0.74 ± 0.04

cytes lysed per neutrophil)

HMPS (cpm ','CO, 11 75 ± 113* 804 ± 83 from (J -"'C)-glu-cose)

Lysozyme release 20.1 ± 2.1 20.7 ± 2.4 (%)

Maximal increase in 7.5 ± 2.9 7.3 ± 1.9 cAMP (pmol 107 cells)

per

Values are mean ± s.e.m. from 12 sepa rate experiment except for cAMP resul ts representing 10 experiments. Monocytes (J X lOf» were incubated with anti-D sensit ized rhesus D-positive erythrocytes (12 X

105) for 30 min. Neutrophils (J X lOf» were incubated with a nti-A

sensit ized type A erythrocytes (6 X 10") for 30 min. HMPS is hexose monophosphate shun t activity.

*: p < 0.05, ": p < 0.02, ": p < 0.01.

106 HERLIN, BORREGAARD, AND KRAGBALLE

• UJ 2.50 f-

~ •

a: ~ 0 2.00

~ •

({)

~ • a:

1.50 ~ • • UJ

• • • •

tOO •

0.50

o ~------;--------r------~--------~---4000 cprn14CO;? 1000 3000 2000

FIG 1. Relation between monocyte ADCC and activity of hexose monophosphate shunt in 12 patients with severe psoriasis. R = 0.8392, p < 0.01. Ratio of sensitized rhesus D-positive erythrocytes to mono­cytes was 12:1, incubation time 30 min.

2.00

..J • • 1E 1.50 0 a: ~ • ~

~ • • • • 0

~ tOO • •

({) •• ~ 8 • a:

~ 0.50 UJ

o+--------T------~r_------T_------_r o 1000 2000 cprn 14C02

FIG 2. Relation between neutrophil ADCC and activ ity of hexose . monophosphate shunt in twelve patients with severe psoriasis. R =

0.7063,.p < 0.02. Ratio of sensitized type A erythrocytes to neutrophils was 6:1 , incubation time 30 min.

both patients and controls (Table I). Cyclic AMP values at the termination of the study were not significantly different from values in normals. In neutrophils, addi tion of sensitized type A erythrocytes a lso caused a normal cAMP accumulation in pso­riasis (Fig 4). In 6 of the patients other phagocytic stimuli were investigated in order to see if a general activation of the oxygen

Vol. 79, No.2

30

~ Qj 25

r--u

\2 Qj 0.

(5 E 0.

a::

~ L 20 « 9 .~

1 U >-

(.)

15

10 L-~~ __ -r ______ r-____ ~ ____ ~r-____ ;-o 2 3 4 5 min

FIC 3. Time course of monocyte cyclic AMP levels during mono­cyte-mediated ADCC. Anti-D sensitized rhesus 0 erythrocytes were added to monocytes at zero time. Closed sym.bols represent va lues from 10 healthy controls and op en symbols represent values from ten patients with psoriasis (mean ± s.e. m.) .

15

~ (jj

r--U

\2 CD 0.

(5 10 E 0 1 0.

a:: :2 « .~ U >-

(.) 5

O ~~r-f--~-------r------~-----+------~-

o 2 3 4 5 min. FIC 4. Time course of neutrophil cAMP levels during neutl'ophil­

mediated ADCC. Addition of sensitized type A erythrocytes to neutro­phils at zero time. Closed symbols represent values obtained from 10 healthy controls ann open symbols represent values from 10 patients with psoriasis (mean ± s.e.m.) .

generating apparatus may exist. Addition of phorbol myristate acetate (PMA), a soluble phagocytic imitator, to monocytes or neutrophils revealed a normal production of superoxide anion in psoriasis (Table 11). Also, complement-mediated neutrophil

Aug. 1982

phagocytosis of serum-treated zymosan par t icles (STZ) resulted in a normal increase in oxygen consumption (Table II) and in normal lysosomal enzym e release (Table II) in psoriatic pa­t ien ts.

S ince binding of IgG-sens it ized erytlU'ocytes through Fc-re­ceptors is required for ADCC this receptor activity was evalu ­ated as the ability of monocytes to form rosettes with sensit ized erythrocytes (Table III). N o diffe rence in rosette formation was observed when erythrocytes were sensit ized wi th the undiluted a nti-D ant iserum also applied in t he ADCC assay.

However , using diluted serum for the sensit ization of eryth­rocytes the percentage of m onocytes forming rosettes fell to lower values in normals than in psoria tics with enhanced mono­cyte ADCC. In 2 patien ts with mild psoriasis « 10% of skin involved) and wit h normal AD CC the fall in rosette percentage during serum dilu t ion was not different from normal (data not ShOWII) .

DISCUSSION

The increased chemotactic, phagocytic, and bactericidal ac­t ivi t ies repor ted in monocytes and neutrophils in psoriasis [1-5] seem to suggest a generalized activa tion of the phagocytes. H owever, the use of differen t chemoat tractants has produced conflicting resul ts [2,3,20,21]' In accordance wit h our previous study [6] pat ients with severe psoriasis showed an enhanced monocyte and neu trophil function assessed as ADCC. T he presen t resul ts suggest t hat enha nced ADCC is caused by increased Fc-receptor activity leading to an increased respira­tory bmst but a normal degranulation dur ing AD CC. Since reduced oxygen radicals generated during t he respiratory burst ar e effector molecules in the ADCC reaction [11,12] the relation between ADCC and respiratory burst is presumably causal.

Following binding of the erythrocyte-ant ibody-complex, bu t preceding the respiratory bmst a nd the degranulation, a tr an ­sient accumula tion of cAMP takes place in monocytes and neutrophils [10]. The normal cAMP accumulation during ADCC in psoriasis together with the previous finding of normal cAMP levels in both unstimulated cells and cells stimulated with isoproterenol, histamine and P GE 1 [6] seem to rule out that abnormal phagocyte activi ty in psoriasis is explained by a bnormal cAMP /cGM P ratio as suggested by oth ers [2-5].

The tra nsient cAMP increment during AD CC does not seem to be related to th e respiratory burst [10] bu t may be related to degranulation caused by complete secretagogues t hat is stimuli provoking r elease of enzymes from both azm ophil and specific

TAB LE II. E ffect of phorbol myristate acetate (PMA) and serum. treated zymosan (S TZ) on m.onocyte and neutrophillnetabolism. in

severe psoriasis

Psoriasis Healthy controls

M O/l.ocytes P MA: 0,- genera tion (fmol! 3.18 ± 0.15 3.54 ± 0.06

cell/ min) Neutrophils P M A: 0 ,- generation (fmol! 12.15 ± 1.00 13.07 ± 0.32

cell / min) 8TZ: O2 consumption (fmol! 4.61 ± 0.37 4.25 ± 0.36

ce LL/ min) Lysozyme release (%) 11.6 ± 1.1 10.8 ± 0.8 Beta-glucu ronidase release 5.7 ± 0.9 5.2 ± 0.6 (%)

Values represen t mean ± s.e. m. from six separate experiments.

CYTOTOXICITY IN SEVERE PSORIASIS 107

granules [22]. T his is compatible with om present findings since degranulation following ADCC was normal whereas ADCC induced HMPS activation was enhanced in leukocytes from psoriatic subj ects. In contrast, ADCC obtained from patients with atopic dermatitis h ave shown decreased cytolysis together with impaired cAMP accum ulation [23] and impaired degran­ulation [24]. Cyclic GMP level during ADCC were not recorded since ADCC will provide no change in cGM P in normal cells [10].

As effector molecules dming monocyte and neutrophil me­diated ADCC oxygen radicals [11,12] have been reported. Ox­ygen radicals are formed dm ing the respiratory burst by reduc­tion of molecular O2 to O2- a reaction which by coupled oxida­tion of NAD(P)H to NAD(P) + is connected to the h exose monophosphate shunt being the ult imate electron donor [25]. Psoriasis patients showed enhanced activation of HMPS that was signfican tly correlated to th e enhanced ADCC. This is in accordance with t he close relationship between ADCC, activa­tion of hexose monophosphate shunt and superoxide anion production found in patients with ciU'onic granulomatous dis­ease [11] and with atopic dermatit is [24]. However, th e presence of hemoglobin in the ADCC assay makes it impossible to m easure cytochrome c reduction and hence superoxide an ion production cannot be recorded during ADCC. Krueger et al. [1] found that in psoriasis basal m onocyte HMPS activity as judged by spontaneous reduction of nitroblue tetrazolium (NBT) was increased after incubation of cells for 48 hI' but not after short time incubation (45 min ). However, in the present study we showed more direct evidence for norm al HMPS activity in unstimulated monocytes and neutrophils in psoriasis measuring 14C0 2 production from (1-1 'IC)- glucose.

In psoriasis, t he enhanced HMPS activity during ADCC could not be expla ined by enhanced activity of enzymes respon­sible for the production of oxygen radicals since the respiratory burst induced by th e soluble phagocytosis simulator and by complement-mediated STZ phagocytosis was normal. The ob­servation that the respiratory burst accompanying Fc-receptor mediated ADCC in psoriasis is enhanced but normal during complem ent-receptor mediated phagocytosis is explained by the increased avidity of monocytes to bind to IgG-sensit ized erythrocytes. Increased Fc-receptor activity has also been de­scribed in sarcoidosis [26] and rheumatoid arthritis [27]. It has been shown experinlentally in guin ea pigs that the expression of Fc-receptor is increased when peritoneal macrophages are activated by mineral oil [28]. Since monocyte ADCC is normal in inactive psoriasis [6] the modulation of Fc-receptor activity may be related to disease activity which is consistent with the findings of the rosette studies. T here is, however, no evidence that this modulation is induced by serum factors [6]. In contrast, Sedgwick, Bergstresser , and H urd [29] recently showed that sera from psoriatics increased zymosan activated superoxide generation in normal granulocytes wh en compared to normal sera.

We are indebted to t he B lood Bank and Blood Grouping Laboratory, Aarhus 1<ommunehospital, Aarhus, for prov iding with the antisera. Mrs. Winnie Heidemann a nd M rs. Jonna Guldberg are thanked for skill ful technical assistance.

REFERENCES 1. Krueger GG, Jederberg WW, Ogden BE, Reese DL: Inflammatory

and immune cell function in psoriasis: II. Monocyte function, lymphokine production. J Invest Dermatol 71:195-201, 1978

TABLE III. The percentage of I'Osetle.(orming I/!onocytes as a function of the degree of el),throcyle sensitization

Psoriasis (n = 5) H ealthy cont rols (n = 5)

Undiluted

87.0 ± 5.1 89.2 ± 1.9

Values a re expressed as mean ± s.e.m .

1:4

78.4 ± 8.5 78.5 ± 7.6

ADCC is ant ibody-dependent cell-mediated cytotoxicity. p -value versus contro l. ': p < 0.05.

Serul1l dilu tion

J:8

79.5 ± 6.0 61.5 ± 7.5

I :W

67.0 ± 5.9' 44.6 ± 4.3

ADCC (erylhrocyt.es lysed

per monocyte)

1.68 ± 0.17 ' 1.16 ± 0.13

108 HERLI N , BORREGAARD, AND KRAGBALLE

2. Wahba A, Cohen HA, Bar-Eli M, GaLlily H: E nhanced chemotactic and phagocytic activities of leukocytes in psoriasis vulgaris. J Invest Dermatol 71:186-188, 1978

3. Wahba A, Cohen HA, Bar-Eli M , Gallily H: Neutropil chemotaxis in psoriasis. Acta Dermatovener (Stockh) 59:441-445, 1979

4. Bar-Eli M , Gallily R, Cohen HA, Wahba A: Monocyte function in psoriasis. J Invest D ermatol 73: 147-149, 1979

5. Sedgwick JB, Bergstresser PR, Hurd E H: Increased gra nulocyte adherence in psoriasis a nd psoriatic ar thri t is. J Invest D ermatol 74 :81-84, 1980

6. H erlin T , KragbaLle K: Enhanced monocyte and neutrophil cyto­toxicity and normal cyclic nucleotide levels in severe psoriasis. Br J DermatoI 105:405-413, 1981

7. Cox JP, Karnovsky ML: The depression of phagocytosis by exog­enous cyclic nucleotides, prostagla ndins, a nd theophylline. J Cell Bioi 59:480-490, 1973

8. R ivkin I, Rosenblatt J , Becker EL: The role of cyclic AM P in the chemotactic responsiveness and spontaneous motility of rabbi t peritoneal neutrophils. J Immunol 115:1126-1134, 1975

9. Sandler TA, GaLlin JT, Vaughan M : E ffects of serotonine, carba­mylcholine and ascorbic acid on leukocyte cyclic GMP and chemotaxis. J Cell Bioi 67:480-484, 1975

10. Herlin T, Kragba lle K: T ransient cycl ic AMP accumulation during antibody-dependent cytotoxicity mediated by monocytes and neutrophils. Immunology 43:771-777, 1981

11. Borregaard N, Kragballe K: Role of oxygen in antibody-dependent cytotoxicity mediated by monocytes a nd neutrophils. J Clin Invest 66:676-683, 1980

12. Katz P, Simone CB, H enkart PA, Fauci AS: Mecha nisms of a nt i­body-dependent cellular cytotoxicity. J Clin Invest 65:55-63, 1980

13. KragbaLle K, E llegaard J , Herlin T : Ant ibody-dependent monocyte­mediated cytotoxicity. Isolation and ident ification by esterase staining of human monocytes detached from monolayers by lowering the temperature. Scand J Haematol 24:399- 404, 1980

14. Kragballe K, E llegaard J , H erlin T : An t ibody-dependent monocyte­mediated cytotoxicity. The effects of adherence and removal of mononuclear phagocytic cells and of in vit ro incubation. Scand J Haematol 24:405-411, 1980

15. Klass HJ , Hopkins J, Neale G, Peters TJ: The estima tion of serum lysozyme: a comparison of four assay methods. Biochem Med 18:52-57, 1977

16. F ishman WH, Kato K, Anstiss CL, Green S: Human serum (3-

0022-202X/82/7902-0 108$02.00/0 TH E JOU R NA L OF I N VESTIGATIVE D E RMATO LOG Y, 78:108-115, 1982 Copyright © 1982 by T he Williams & Wilkins Co.

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glucuronidase: Its measurement and some of its properties. Clin Chim Acta 15:435-447, 1967

17. Herlin T , KragbaLle K: Impaired monocyt e cyclic AMP responses and monocyte cytotoxicity in atopic dermatit is. Allergy 35:647-655, 1980

18. Borregaard N, Juhl H: Activation of the glycogenolytic cascade in human polymorphonuclear leukocytes by di fferent phagocytic stimuli . Eur J Clin Invest 11:257-263, 198)

19. Borregaard N, J ohansen KS, Esmann V: Quant itation of super oxide product ion from normals and 3 types of chronic granulomatous disease. Biochem Biophys Res Commun 90:214-219, 1979

20. Krueger GG, Hill HR, Jederberg WW: Inflammatory and immune cell function in psoriasis-a subtle disorder I . In vivo and in vit ro survey. J Invest Dermatol 71 :189-194, 1978

21. Breathnach SM, Carrington P , Black MM: Neutrophil leukocyte migration in psoriasis vulgaris. J Invest D ermatol 76:271-274, 1981

22. Smolen JE, Weissma nn G: S timuli which provide secretion of azurophil enzymes from huma n neutrophils induce increments in adenosine cyclic 3'-5'-monophosphate. Biochim Biophys Acta 672:197- 206, 1981

23. Kragballe K, Herlin T : Ant ibody-dependent monocyte- mediated cytotoxicity in atopic derma titis. R elation to clinical s tate a nd cyclic AMP response. Allergy 36:27-32, 1981

24. Kragballe K, Borregaard N: Mechanisms of decreased a ntibody­dependent cytotoxicity media ted by monocytes and neutrophils in atopic dermatitis. Acta Dermatovener (Stockh) 61:11- 15, 1981

25. Babior BM: Oxygen-dependent microbial killing by phagocytes. N Engl J Med 298:659-668, 1978

26. D ouglas SD , Daughaday CC, Schmidt ME , Siltzbach LE: Kinetics of monocyte receptor activity for immunoproteins in patients wi th sarcoidosis. Ann NY Acad Sci 278:190-200, 1976

27. K atayam a A, Chia D, Nasu H, Knutson DW: Increased Fc recepto r ac tivity in monocytes from patien ts with rheumatoid ar thrit is: A study of moncyte binding and catabolism of soluble aggregates of IgG in vi tro. J Immunol 127:643-647, 1981

28. Rhodes J : Macrophage heterogeneity in receptor activity: The activa tion of macrophage Fc recepto r function in vivo and in vitro. J Immunol 114 :976-981, 1975

29. Sedb'Wick JB, Bergstresser PR, Hurd EH: Increased superoxide generation by normal granulocytes incubated in sera from pa­tients with psoriasis. J Invest Dermatol 76:158-163, 1981

Vol. 79, No.2 Printed i/l. U.S.A.

Mechanisms of Contact Photosensitivity in Mice: I. T Cell Regulation of Contact :photosensitivity to Tetrachlorosalicylanilide under the Genetic

Restrictions of the Major Histocompatibility Complex

MASAHIRO TAKIGAWA, M.D., AND YOSHIKI MIYACHI, M .D .

Department of Dermatology, Faculty of M edicine, Kyoto University, Kyoto, J ap an

Induction, transfer and specificity of contact photosen­sitivity to 3,3' ,4' ,5-tetrachlorosalicylanilide (TCSA) w ere s tudied in mice. Mice were sensitized by 2 daily abdominal paintings with 1% TCSA plus irradiations with "black lights." The degree of hyper sen sitivity was a ssayed by m ea suring the increment of ear thickness 24

Ma nuscript received November 10, 1981; accepted for pu blication January 18, 1982.

Reprin t requests to: Dr. Masahiro T akigawa, Depar tment of Der­matology, Facul ty of Medicine, Kyoto University, Kyoto 606 J apan.

Abbreviations: BITH: bithionol C: complement DNFB: dinitroflu orobenzene DTH: delayed-type hypersensitivity HBSS: Hanks' Bala nced Salt Solu tion

hr after challenge with 0.1% TCSA plus irradiation with black lights. Long ultraviolet (320-400 nm) but not ery­throgenic (280-320 nm) radiation was required for in­duction and elicitation of the response. The 24-hr ear reaction was not detectable on day 3, peaked on days 5 to 7, and waned thereafter. Tpe histologic picture of a swollen ear at the peak of the reaction showed dense infiltration of mononuclear cells intermingled with pol-

I

MHC: major histocompa tibility complex NP: 4-hydroxy-3-ni trophenyl acetyl OX: oxazolone TBS: 3,4',5- tribromosalicyla nilide T CS: 3' ,4 ',5-trichlorosalicylanilide . T CSA: 3,3',4' ,5-tetrachlorosalicyla nilide UV A: long ultraviolet irradiation UVB: erythrogenic ultraviolet irradiation