Characteristics of prostaglandin E1 potentiation of inflammatory activity of some agents

PROSTAGLANDINS

CY~ACTERISTICS OF PROSTAGLANDIN E 1 ~IATION OF ~ R Y

~CrIVITY OF SOME AG~WPS

GEORGE THCMAS

Department of Pharmacology &Tnerapeutics

University of Ibadan, Ibadan, Nigeri~

Present address:

Laboratory of Pharmaceutical Technology,

Federal University of Paraiba,

58.000 - ~ PESSOA - PARAIBA - BRAZIL.

ABSTRACT

In rats pretreated with ~thacin, injection of PGE I (prostaglandin E l) with carrageenan potentiated the carrageenan paw oedema. This effect of PGE~, was maximal when it was injected together with carrageenan, there being a reduction in the action of PGE 1 if carrageenan injection was delayed after PGE 1 injection. PGE 1 induced potentiation of increase in plamma protein leakage induced by intradermal injections of bradykinin and histamine also depended on the injection of PGE 1 along with these agents, qhus oedema enhancement by PGE 1 differs ~rcm its action in pain, where PGs cause a long lasting sensitlzation of the injected area for the actions of other algesics. Since vasodilation may be a mechanism of oedema enhancement by PGs, the ability of adenosine and papaverine to mimic PGE 1 in paws and skins of rats were exanined. Adenosine was active whereas papaverine was inactive in this respect. To clarify this difference, the vasodilatory properties of PGE I, adenosine and papaverine were assessed by their ability to antagon/~e NA re~onse in perfused rat mesenteric blood vessels. Only papaverine was effective in antagoni- sing the NA response. Tnus, PGE 1 and adenosine which potentiated the oedema inducing actions of other agents showed no vasodilatory properties and papaverine, a vasodilator, had no oedema potentiating actions.

JANUARY 1980 VOL. 19 NO. 1 39

PROSTAGLANDINS

~ C N

Simultaneous ac~ninistration of prostaglandin E 1 (PGE l) , enhances the increase in vascular permeobility induced by other pro-inflam~tcry agents. Thus, it potentiates the carrageenan response in the rat paw (1,2), potentiates the plasma protein leakage caused by bradykinin in the rat skin (3) and by bradykinin and histamine in the dog kneejoint (4). However, few attempts have been made tour~stand the characte- r/stics of such potentiating actions of PGE I. In the present experiments with rats using carrageenan paw cedema and increased plasna protein exudation caused by intradermal injection of bradykinin, histanine and 5-hydrox~tanine (5-HT) attempts were made to study the time course of PGE 1 potentiation. Further, vasodilatation which has been proposed as a possible ~ ~ of PGEI potentiation (5) was studied by investigating the ability of two vasodilators, adenosine and papaverine to mimic the action of PC~ 1 in paw and skin experiments in the rat. Since adenosine and papaverine gave different results, preli- minary studies were also made regarding the vasodilatory properties of these agents and P(~I using rat isolated mesemteric blood vessel preparations (6).

METHODS

Time course of ~otentiaticn of carrageeman paw oedema with. IK~

Male Wistar rats (120 - 150g) in groups of 7 were given 20 mg kg -I, p.o. of indnmethacin, 30 minutes before the injection of carrageenan (i mg/paw) into the right paws, to suppress endogenous PG producticm. At 60, 30, 15 and0min beforecarrageenan, P~ in doses of 0.I, 0.5 or 2.5 ~g/paw were also injected into the right p~ws. For comparison, other groups of indnmethacin treated rats received either carrageenan or Iq~ 1 alone. Rats were killed one hour after carrageenan, and the paws removed at the tibio-tarsal junction, weighed and the weights of the right paws were crmpared with the weights of saline injected left paws. To assess the potentiating effect of IK~I, the weigts of the paws given both PGE 1 and carrageenan were compared with those receiving carrageenan alone and the results analysed statistically using student's 't' test. To calcula- %m The extent of potentiation by IK~ 1 of the carrageenan response, the results ob~alned with the agents when given alone were added and the total was subtracted from the results obtained when the two agents were given together. The differences were expressed as percentages of the totals of the responses when the agents were given alone.

Potentiation of Carragee~an ~.w oedema with adenosine and papaverine

The met/xx~ %~s similar to that described above for I~ I except t-hat adenosine and papaverine was always given along with carragee~an. The dose of adenosine ranged from 1 - i00 ~g/paw while that of

40 JANUARY 1980 VOL. 19 NO. 1

PROSTAGLANDINS

papaveriDe was from 1 - 20 ~g/paw. Higher doses of papaverine were not tried as the drug precipitates at higher concentrations at ~hysiological pH values.

Time course of PGE I potentiation of bradykinin, histamine and 5-HT

responses in the rat skin

Male Wistar rats (150 - 200 g) ingroul~S of 6were anaesthetised with ether and 0.i m!/100 g body weight of a 20 mg/ml solution of Evans blue was injected into the tail vein. This was followed by intradermal injection of PGE 1 ( 1 - i00 ng/site) into previously depilated skin on the back of the animals. At 60, 30, 15 or 0 minutes after PGEI, threshold doses of bradykinin (i00 ng/site), histawhne (500 ng/slte) or 5-HT (i0 rig/site) were injected at the site of previous PGE 1 injections. For comparison, PGE 1 ca= the inflammtozy agents were injected alone as well. All the drugs were dissolved in sterile Tyrode solution and the volune injected at any one time was 0.05 mls. An hour after the injections of bradykinin, histamine or 5-HT, the animals were k111ed and the skin removed. The dye exuded at the sites of injections was extracted and assayed spectro~to- metrically as described by Harada and others (7) .To test the effect of PGE I, values obtained when both PGE 1 and the inflammatory agent were LSjected together were cc~pared w~th the values obtained when the inflam~tory agent was given alone and the results were analysed statistically using student's 't' test. To detennine the extent of PGE 1 potentiation, the values obtained with PGE 1 and the inflammatory agent when given alone were added and the total was subtracted from the results obtained when the two agents were given together. The differemces were expressed as percentages of the totals of the response when the agents were given alone.

Action of adenosine and ~a~verine on brad~kinin t h/stanlne and 5-HT responses in .~_ rat skln

The procedure was similar to that enloloyed above for PGE 1 ~xceot that both adenosine and papaverine were injected together with the inflam~tcry agents. The doses of adenosine and papaverine ranged from 0.i - i0 pg/site.

C~ison of the vasodilator ~ actions of PGEI, ' adenosine and papaverine

Vasodilatory action of the drugs was measured by using the isolated perfused rat mesenteric blood Vessels (6). The activity was assessed by the ability of the drugs to inhibit the rise in perfusion pressure (ram Hg) produced by injections of noradrenaline (NA) in 100-500 ng doses. After obtaining a suhmaximal response (about 50% of the maximal response) to NA, the tissue was perfused with the test


PROSTAGLANDINS

drug.Two minutes after, the dose of NA was again repe2ted and the increase in pressure was recorded using a Devices 4 Channel recorder. The response to NA after the drug was ccr~pared with the control NA response and expressed as a per cent of the control response. At least 4 preparations were used for each drug.

RESULTS

Time course of potentiation of carrageenan paw oedema with PGE 1

Figure IA & B, demonstrates the increase in paw weight produced by PGE I. alone, carrageenan alone, or when carrageenan, was given at various intervals after the injections of PGE 1 into the paws. PGE 1 response was measured one hour after the injection and in all the other cases, the responses were those ~b~ained one hour after the carrageenan injections. Figure IA shows t-he potentiating action of PGE 1 at 0.1~/paw and in Figure IB, the effect of 0.5 ~g/paw of P~l is given. O3mpared with the saline response (not shown) the smaller dose of P~I failed to cause a significant (P > 0.I) increase in paw weight. However, PGE~ at 0.5 ~g/p~. and carrageenan at 1 rag/paw always produced a signiflcant effect (P < 0.01). Simultaneous injections of PGE 1 in either 0.i ~g or 0.5 ~g/paw doses with carrageem~n produced responses which were significantly greater (P < 0.001) than the responses to carrageenan alone. ~he effects of combined injections ofPGE 1 either in 0.1 ~g or 0.5 ~g doses with carrageenan were greater titan the sum of the effects of the individual drugs given alone, by 59% and 74% respectively. Results also show that if carrageenan injection was delayed 15, 30 or 60 minutes after PGE 1 injection there was a corresponding reduction in the degree of potentiation of the carrageer~n respcr~e. For exaa~le, while carrageenan adninistration, 15 minutes after the injection of PGE 1 0.i ~g/pa~ (Figure IA) produced a response which was significantly greater (P < 0.01) than the response to carrageenan alone, the increase in paw weight (600 rag) due to the cunbination of drugs was only 25% greater than the sun of the individual drug effects (480 mg). Similarly even though the injection of carrageenan, 15 min. after PQ~I 0.5 ~g/paw (Figure IB) gave a significant (P <0.05) increase ~paw weight (580 mg) when compared with the control carrageeD~n response, the increase was only 8.4% greater than the sun of the the responses (535mg) to PGE 1 and carrag~ when given separately.

Increasing tb~ dose of PGE 1 to 2.5 ~g/paw (not shown) did not increase the carrageenan response any further than 0.5 ~g/paw dose and the potentiating effect of the higher dose of PGE 1 also depended on its injection along with the carrageenan injection.


P R O S T A G L A N D I N S

Figure 1 Time oourse of potentiation of carrageenan paw oedema with PGE 1

(A)

(B)

Shows the mean increase in paw weight in grams (± s.e.m.) produced by the injections of 0.1 ~g/paw of l~l alone, carrageenan alone or when carrageeman was injecuea at 0, 15, 30 or 60 minutes after the injections of PGE 1 into the paws. PG = Prostaglandin . C = Carragsenan

Gives tb~ results obtained when a higher dose of PGE 1 (0.5 ~g/ paw)was used. In both 0~) and (B), P~E 1 response %ms measured one hour after its injections and in all the other cases, the responses were assessed c~e hour after the carrageenan injections.

A PGE10"I~g

I I"

a .

1"

Inn C . . . . a .

, i

O. a . a . a .

1" B PGE10,5~g

I

T

i

C e- e- g~ , ~ i m , i

a . f t . a . a .


PROSTAGLANDINS

Potentiation of carra~eex~n ~ oedema with adenosine and papaverL~

Table 1 shows the results for adenosine and papaverine when compared with PGE 1 in enhancing the carrageenan response in indcme- thacin pretreated rats. When injected alone, both adenosine and papaverine failed to produce a significant increase in paw weight compared to saline injected paws. Simultaneous injection of adenosine (50 ~g/paw) with carrageenan enhanced the carrageenan response in a significant manner (P<0.01). However, the effect was far less than that produced by PGE 1 in 0.5 pg/paw dose. Thus, while the effect of combined injection of IK~ 1 and carrageenan was greater than the sum of the effects of the individual agents by 70.9%, the result for aaenosine and carrageenan was 30.0%. Smaller doses of adenosine were not effective and the effect of a higher dose (i00 ~g/paw) of adenosine was not different from that of 50 ~g/paw dose. Papaverine at 20 ~g/paw, which was the highest dose that was possible had no significant effect on the carrageenan response. Tne combined injection of these two agents produced an effect which was lower than the sum of the responses of the agents when given separately by 23.7%.

Time course of PGE 1 potentiation of bradykinin, histanine aD~ 5-HT

responses in the rat skin . . . . . . . . .

PGE 1 in a range of doses (i - i00 ng/site) potentiated the responses to intradermal injections of threshold doses of bradykinin (i00 ng/site), histamine (500 ng/site) but not to that of 5-HT (10 ng/site) into the rats. As an example,while PGE 1 (i0 ng) induced 2.0 ± 0.4 micrograms and bradykinin, 6.0 ± ~.6 micrograms of pla~na protein bound dye leakage, the combined injection produced 16.5 ~ 1.0 micrograms of dye exudation, which was significantly (P < 0.001) greater than the effect of bradykinin alone. The combined response was greater than the sun of the individual responses to the two agents (8.0 microgranes) by 106.0%. If the bradykinin injection was delayed after PGE 1 injection, there was a progressive reduction in enhancenent of bra~ykinin response and no significant potentiation occurred if the time interval between the two injectior~ was greater than 30 minutes. Thus, if bradykinin was injected 60 minutes after PGE 1 it produced exudation of 9.0 + 1.0 micrograms of dye, the results for the separate injection of the agents being 6..3 +_ 0.8 and 1.8 + 0.3 microgram of dye for bradykinin and PGEI, respectively. Tne man of the effects of the two agents (8.1) wh~% given separately was not different from the effect of combined injection (9.0) of the two agents.

Similarly, maximun enhancement of histamine response by PGE 1 also depended on the injection of PGE 1 along with histamine. P(~I did not potentiate 5-HT responses.


P R O S T A G L A N D I N S

~ I~ ~ ~

• "4 t".O

I + I + 14- I +

I ~ I-~ t.O I'O

0 0

_~ ~ ~ ~ o~ ~

&

o , ~ ~

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~ ' ~ . ~

, 00 ~'~"

!

• . ~ I+ I + I + I + I + +

• ~ ' ~

N.~ P'~ t.O t.O ",4 ',,O O O -.4 O +


PROSTAGLANDINS

Action of adenosine and papaverine on bradyklnin, histanine and

5-HT r egloonses in the rat skin.

Table 2 demonstrates the action of adenosine in potentiating the increase in vascular permeability produced by bradykinin, histamine and 5-HT after intradermal injections in the rat. Adenosine when injected along with the agents in doses of 1.0 and i0 ~g/site produced a significant (P < 0.01) potentiation of these agents. Results with 0.I ~g/site of ader~sine were not significant. Higher doses of adenosine were not tried as these doses produced significant dye leakage cn t_helr own. Adenosine was less effective in potentiating 5-HT responses than either bradykinln or histanine responses.

Results obtained with papaverine (i0 pg/site) are given in Table 3. Papaverine failed to enhance the actions of bradykinin, histamine and 5-HT in the rat skin. Ccmbined injections of papaverine with each one of the above agents produced responses which were not significantly different frcm the effects of these agents when given alone. In fact, the effect of combined injections of the above agents with papaverine always produced responses which were smaller than the sun of the effects of the agents and papaverine when they were given separately. However, these effects were not significant (P • 0.I). Smaller doses of papaverine were also ineffective.

Comparative vasodila~ action of PGE I, adenosine and papaverine

in mesenteric blood ~essels.

Injections of either PGE I or adenosine into the fluid perfusing the tissue (in doses given in Table 4) had no effect on the mean resting perfusion pressure of 20 - 25 nm .Hg. However, papaverine always caused a reducticm in perfusion pressure, but this effct could not be quantified as it w~s of s~%ll magnitude as the preparation e~h/bits little resting tone.

Effects of the agents on NA induced increase in perfusion pressure are also given in Table 4. It shc~s that PGE 1 in 0.I ~g ~nd adenosine in i0 and 50 ~g doses significantly (P < 0.01) er~-mnc:~, t he NA responses. On the contrary, papaverlne in 10 and 20 ~g doses caused a significant (P < 0.01) dose related inhibition of NA induced rise in perfusion pressure.


PROSTAGLANDINS

Table 2

Effects of various intradenmg/ doses of adenosine on dye leakage produced by intradenTal injections of bradykinln, histamine and 5 - h y ~ in doses of i00, 500 and i0 ng/site respectively into the rats. Results are mean + s.e.m, of dye (~g) extracted fram the skin of groups of 7 rats one hour after the injecticms.

Agent under test

Agent injected Interaction of the agent with adenosine

0.i (~/%ite) i0.0

Bradykinin alone (a)

i Adenosine alone (b)

Sun of (a) + (b)

m Cnmbined injection of a and b (c)

% Potentiation

6.8 + 0.5 6.8 + 0.5 6.8 + 0.5

3.6 + 0.3 3.0 + 0.4 6.5 + 0.5

10.4 9.8 13.3

12.2 + 0.7 17.6 + 1.3" 27.4 -+ 3.0*

+ 17.3 + 79.6 + 106.0

Histamine alone (a) 4.7 -+ 0.5 4.7 + 0.5 4.7 + 0.5

Adenosine alone (b) 3.2 + 0.4 2.7 + 0.3 7.0 + 0.9

Sum of (a) + (b) 7.9 7.4 11.7

Combined injection of a and b (c) 9.8 + 0.8 13.5 ± 1.3" 22.3 + 3.6*

% Potentiation + 24.0 + 82.4 + 99.1

u%

5-h'f alone (a) 5.2 +- 0.6 5.2 + 0.6 5.2 + 0.5

Adenosine alone (b) 3.1 ± 0.2 3.1 + 0.3 5.5 ± 0.7

Sum of (a) + (b) 8.3 8.3 10.7

Ccn%bined injection of a and b (c) 8.2 + 0.9 12.4 + 1.0" 17.1 + 1.0"

% Potentiation - 1.2 + 49.5 + 60.0

* Significant at P < 0.01 % Potentiation = C- (a +b) x 100

(a + b)


PROSTAGLANDINS

Table 3

Effect of papaverine (i0 ~g/site) on dye leakage induced by intradermal injecticms of bradykinin (i00 ng/site), histamine (500 ng/site) and 5-HT (i0 ng/site) into the rats. Results are mean + s.e.m, of microgran of dye exuded/site one hour after the injections in groups of 5 rats. (bk. = Bradykinin; Hist. = Histamine).

Agent Dye leakage Dye leakage by Dye leakage by % Potentia- tested by agent papaverine combination tion

(a) (b) (c) C- (a+b) x 100

(a+b)

Bk. 8.2 ~ 0.7 3.8 ± 0.9 9.2 ± 2.2 - 23.3

~t. 8.8±0.8 " 8.9±1.0 - 16.0

5-HT 7.0±1.1 " 8.3 ~1.2 - 23.1

Table 4

Effects of PGEI, adenosine and papaverine on NA induced J_ncrease in perfusion pressure in rat isolated mesenteric blood vessels. Results are either an enhancement (+) or inhibition (-) of oontrol NA responses when the agents were administered 2 minutes before NA injection. * = Significant at P < 0.01 when the NA responses before and after the drugs were oompared.

Drug Dose % effect on NA induced responses (~g) (Mean + s.e.m.)

I o.ool + lO.O +_ 2.1 0.01 + 53.1 + 4.0*

Adenosine i0 + 21.4 + 3.6* " 50 + 62.3 + 7.0*

Papa,v, erine 5 - 16.0 + 2.2 i0 - 39.7 + 6.1"

" 20 - 50.5 +- 6.8*

DISOJSSIC~

Results show that in the rats predosed with indomethacin to suppress endogenous PG synthesis, injections of PGE 1 in doses of 0.i, 0.5 and 2.5 ~g/paw along with carrageenan into the paws potentiated the carrageenan oedema. The action of PGE 1 may be due to its potentiation of the effects of histamine and 5 hydroxy- tryptanine as the carrageenan paw oedema in the first hour is mediated by the release of these two agents (8). The potentiation depended on the sin~itaneous presence of PG along with carrageenan, there being a fall in the potentiating effect of PGEI, if the carra~- eenan was delayed after PGE 1 injection. Similarly intrade2m~l experiments in the rat also demoz~trate that for PGE 1 induced potenti- t~on of plasma protein leakage produced by br~ykinin and his tanine


PROSTAGLANDINS

PGEI has to be injected along with these agents. %'nus, for maxamal oedema potentiating activity, PGE 1 has to be present at the time of injection of the pro-inflammatory agent. This is unlike its action in pain production, where on injection PGs cause a long lasting sensitization of the injected area for the action of other algesic agents such as bradykinin and histamine (9).

It has been proposed by Williams and Peck (5) that in the rabbit PGs enhance the oedema producing actions of other media- tors by their vasodilatory actions and thereby increasing the local blood flow. One piece of evidence was that adenosine, a vasodilator in the rabbit, mimicked the PGE 1 potentiaticm. In an atte~ot to extend these observations to the rat, the potentiating effects of PGE 1 were ccmpared with adenosine and another vasodilator, papaverine in the paw and skin experiments. In carrageenan paw oedema results indicate that PGE 1 in 0.5 ~g/paw dose produced a greater enhancement of the response than a dose of 50 ~g/paw of adenosine and papaverine in a dose of 20 ~g/paw had no such effect. In the intrademmal experiments adenosine (in doses of 0.i - i0 ~g/site) was able to enhance the effects of bradykinin and histanine and further, unlike PGEI, adenosine alsoer~anoed the 5-HT response as well. Papaverine on the other hand was again inactive in doses up to i0 ug/site.

In order to clarify the different results obtained with PGEI, adenosine and papaverine, their relative vasodilatory properties were assessed using the isolated perfused rat mesenteric blood vessels in which vasodilaticn is measured by the ability of the agents to antagonize the rise in perfusicn pressure produced by NA, a vasoconstrictor. It was found that while papaverine antagcnized the NA iDzluoed rise in perfusion pressure, PGE 1 and adenosine potentiated the NA effect. Other evidenee is also available to suggest that PGE 1 and adenosine have vasooonstrictor actions. For example, Chahl and Ladd (10) showed that PGE 1 in i0 - 200 ng/ml doses produced constriction of rat cutaD~ous blood vessels, and adenosine had been proposed as a mediator of hypoxic renal vasoconstrictor in the dog and the rat (ii). Further, adenosine potentiates.renal vaso- constriction induced by ~thetic nerve stimulation in the rabbit (12).

Thus if the present results with the mesenteric blood vessels and those results reported by others (i0, Ii, 12) can be extended to the rat cutaneous vasculature, they suggest that PGE 1 and adenosine have no vasodilatory actions in the rat skin and that the oedema potentiating effects of the two agents are not due to their vasodilatory properties. This is further supported by the inability of papaverine, a vasodilator, to enhance the inflammatozy actions of carrageenan, bradykinin, histamine and 5-HT in the present experiments. However, oonfimmation of the above oonclusions can only be


PROSTAGLANDINS

obtained by the slaE%ltaneous measurements of the effects of the agent on exudation potentiaticm and blood flow changes in the sa~e tissues of the rats.

ACKN~L~DG~V2S

I wish to thank Mr. A.S.O. Adeagbo for performing the experiments cn rat mesenteric preparations, Dr. J. Pike for the gift of PGE 1 and Dr. H. Friedli for the gift of bradykin/n.

i. Monc~,S., Ferreira, S.H., and Vane, J.R., Nature, 246, 217 (1973).

2. Lewis, A.J., Nelson, D.J., and Sugrue, M.F., Br. J. Pharmc., 55, 51 (1975).

3. Tncmas, G. and West, G.B., ~r. J. Phannac., 50, 231, (1974).

4. Grennan, D.M., Mitchell, W., Willer, W. and Zeitlin, I.J., Br. J. Phannac., 60, 251, (1977).

5. Williams, T.J., and Peck, M.J., Nature , 270, 530 (1977).

6. McGregor, D.D., J. Physi01., 177, 21 (1965).

7. Harada, M., Takeuhi, M., Fukao, T. and Katagiri, K., J. Phann. Phazmac., 23, 218 (1971).

8. Di Rosa M.,Giroud, J.P. and Willoughby, D.A.J. Path., 104, 15 (1971).

9. Ferreira, S.H., Moncada, S. and Vane, J.R. In Prosta@landin Synthetase Inhibitors, Robinson, H.J. and Vane, J.R. Eds., Ravem Press, New York, P. 175 (1974).

i0. Chahl, L.A. and Iadd, R.J., B_{r. J. Pharmac., 56, 317, (1976).

ii. Fredholm, B.B. and Hedqvist, P., Br. J. Pharmac.,64, 239 (1978).

12. Hedqvist, P. and Fredholm, B.B., Naunyn-Schniedeber~'s Arch. Pharmac., 293, 217 (1976).

Editor: F. Kuehl Accepted 11/22/79


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Characteristics of prostaglandin E1 potentiation of inflammatory activity of some agents

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