THROMBOSIS RESEARCH 16: 329-333 Pergamon Press Ltd.1973. Printed in Great Britain

EFFECT OF HRPARIN ON PULMONARY EDEMA AFTER MICROEMBOLIZATION IN RATS*

Arnold Johneon and Asrar B. Malikt Departmant of Physiology Albany Medical College of Union University

Albany, New York 12208

(Received 29.5.1979. Accepted by Editor P.C. Harpel)

ABSTRACT Similar degree of pulmonary microembolization was induced in non- heparlnieed and heparinised rats by injecting 50 WI diameter glass beads into the jugular vein. At 1 hour after the embolisation the extravascular lung water content/bloodless dry lung wefght in the non-heparinized was increased from control value of 3.32i.067 to 13.06k2.32 (p&OS); but the increase (~~0.05) in the heparinieed rats of 7.532.48 was less (p&.05) than in the non-heparinized rats. Fibrin accumulated in the non-heparinized rata after emboli- matlon, but not in the heparlnized animals. The results suggest that release or activation factors subsequent to the fibrin accumulation induced by pulmonary microembolization attenuates the development of pulmonary edema in the rat. Since heparin pre- treatment in rats did not completely prevent the pulmonary edema as in dogs, factors in addition to those associated with fibrin accumulation also contribute to the fluid accumulation in the rat lungs after microembolization. The reaults suggest that the development of edema after pulmonary microembollsation cannot be attributed only to fibrin accumulation in all species, but other factors may also be involved.

INTRODUCYION

pulmonary microembolism syndrome is characterized by intravascular co- agulation, platelet aggregation and leukocyte margination in the pulmonary vessels (1). Humoral factors, which are activated and released after pulmonary

*Supported by NIH Grant IU-17355 and Research Career Development Award to A.B. Malik (HL-00363). A.Johnson is a doctoral trainee supported by HL-07194.

ITo whom reprint requests should be addressed: Department of Physiology, Albany Medical College of Union University, 47 New Scotland Avenue, Albany, New York 12208.

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microembolism may play a role in the development of pulmonary edema and pulmonary insufficiency (1) associated with microembolism. Fibrin and its degradation products (FDP) have been Implicated as the edemagenic factors in the lungs (1,2,3). Hallk and van der gee (2) inhibited the increases in extravascular lung water following pulmonary microembolization in dogs with heparln pretreatment prior to the glass bead microembolization suggeeting a role of secondary thrombosis in the genesis of pulmonary edema. Inhibition of fibrinolysis in dogs with tranexamic acid resulted in increased fibrin uptake which was aseoclated with increased extravascular lung water content (3) suggesting a role for the fibrin entrapment in the development of pulmon- ary edema after microembolism. These previous studies have been in dogs; it is unclear whether this effect is specie8 specific because of differences in the degree of intravascular coagulation (4). In sheep heparin pretreatment and defibrinogenatlon did rwt prevent the increase in lung vascular permea- bility that results after pulxonary embolization suggesting that fibrin and its degradation products do not play a role in the development of lung vascular injury (5). The purpose of this study was to determine the effect of heparin pretreatment on lung extravascular water accumulation after micro- embolization in rats and to c-are the effects with our previous studies in dogs (2,3).

METHODS

Sprague-Dawley rats (460 f 20 g) were anesthetized with an intraperi- toneal Injection of sodium peutobarbital (25 mglkg). The left jugular vein and right carotid were catheterized. Tracheotomies were made and the rats were were ventilated with a small animal respirator (Phlpps and Bird, Blchmoud, VA) to establish normal blood gases and pH at the beginning of the experiment Non-slliconized glass beads approxieretely 50 um in diarPeter were suspended in 1 to 2 ml saline, and were infused into the jugular vein over a 5 min period.

Studies were made on three groups of rats;. Group I (n = 5), rats with- out bead infusion but receiving the saline; Group II (n = 5), rats infused with beads; aud Group III (n = 5), rats pretreated with heparin (500 u/kg) one hour before the sope degree of emboliaation as Group II. All animals were killed one hour after the embolization.

The extravascular lung water content/bloodless dry lung weight ratio was determined at the end of the study by the method of Pearce (6) with the correction made for the weight of beads that were injected (2). The following equations were used to compute the extravascular lung water content and blood- less dry lung weight:

Bxtravascular lung water content = wet weight - lung blood weight - bloodless dry lung weight - bead weight.

Bloodless dry lung weight - dry lung weight - weight of formed elements in the lung - bead weight.

The residual blood content in the lung was determined from the hemoglobin concentration in the lung homogenate of the rehydrated lung (5).

Fibrin accumulation in the lung after microembolization was determined by injecting 1251-fibrinogen (100 uCi) one hour prior to the experiment

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(7). An arterial blood sample was taken at the end of the experiment for determination of 1251 activity per g blood. The lung and blood activities were counted in a Nuclear-Chicago gam counter. The amount of fibrin trapped in the embolized lungs was assessed by the ratio of total lung activity to the expected lung activity. The expected lung activity was obtained from the lung blood weight which was determined as described above (6).

The means were compared using the Student's t-test setting the con- fidence level at 95%.

BBSDLTS

The data in Table 1 indicate significant increases (pcO.05) in extra- vascular lung water (BLW) and the extravascular lung water content/bloodless dry lung weight ratio (ELW/BDL) after embolization (Group II) as compared to the control non-embolised rats (Group I). Gross tracheal edema was observed in all embolixed rats 1 hour after the embolixation. The untreated embolieed rats also had significantly less (~~0.05) blood trapped in the lungs than control rats.

Table 1. Effects of pulmonary microembolization and heparin treatment followed by microembolization on the bloodless dry lung weight, residual lung blood volume, extravascular lung water content, and extravascular lung water content/bloodless dry lung weight ratio. The values are indicated on mean f ISEM.

Bloodless Dry Lung Wt. (g)

CONTROL (n = 5)

.39+.10

MICEGEMBOLI (n - 5)

.29? .070

HEPABIN + MICEGEMBGLI

(n = 5)

.30+.038

Lung Blood Volume (ml)

.87+-.24 .24+ .05* .31+.013*

Extravascular Lung Water (ml)

1.32k.35 3.43* .70* 2.18+.20*t

Extravascular Lung Water (ml) 3.322.067 13.06+2.32* 7.53+-.48*t Bloodless Dry Lung g

* - different from control (~~0.05) t = difference from untreated, embolized (~~0.05)

The heparin pretreated rats (Group III) also had greater increases (~~0.05) in ELW/BDL than controls, but ELW and ELW/BDL were significantly less (~~0.05) than in the untreated embolized rats. Fulminant airway edema was not observed in this group as in the untreated group. The heparin group had significantly less (~~0.05) blood trapped in the lungs than control rats and the value was not different from the untreated embolized rats. The total

332 HEPARIN AND PULMONARY EDEMA

1251-activity/expected 1251-activity ratio of 3.62kO.44 in the heparinized group was less (~~0.05) than the value of 18.3427.92 in the non-heparinized group indicating significant fibrin accdation in the untreated embolized group. The value in the heparinixed rats was not different from the control value of 2,18x0.51.

DISCUSSION

The results indicate that heparin pretreatment in rats attenuated the increase in extravascular lung water content after pulmonary embolization. These data are consistent with our previous findings in dogs indicating the salutary effect of heparin pretreatment in inhibiting the increase in extra- vascular lung water after pulmonary embolization (2,3). However the results indicate quantitative differences between rats and dogs. In the present study in rats, heparin pretreatment did not prevent the microembolixation- induced pulmonary edema; but rather the degree of fluid accumulation in the lungs was diminished. Although there are these quantitative differences between rats and dogs, the data support the hypothesis that fibrin entrapment in the lung subsequent to pulmonary microembolism contributes to the genesis of pulmonary edema in both species (8).

There may be several reasons why heparin did not prevent the pulmonary edema after pulmonary microembolization in rats as it did in dogs. It is possible that the pulmonary edema in heparinized rats may be due to differences in the degree of embolixation in heparinized and non-heparinized groups. How- ever the differences in the degree of lung water acc=lation in heparinized and non-heparinixed rats cannot be explained by differences in the degree of embolixation since the same number of beads were injected in both groups and the extent of pulmonary vascular obstruction was comparable as evidenced by similar decreases in the lung blood volume in both embolized groups. The presence of pulmonary edema in the heparin pretreated rats may thus be due to the release or activation of humoral factors which are independent of fibrin accumulation in the lung. There are several possibilities. Pulmcnary micro- embolization in the heparin pretreated rats may result in the release of histamine via the mast cell degranulation which may in turn increase lung vascular permeability and contribute to pulmonary edema (9). Prostaglandins are released from embolized lungs (9) and hence these may also be involved in the genesis of edema. Finally, the entrapment of platelets and leukocytes in lungs after embolism and the subsequent release of lysozymes as well as the activation of the kallekrein-kinin and complement systems as the result of Factor XII activation after glass bead embolization may play a role in mediating the pulmonary edema in the heparinized rats. Thus other humoral factors associated with fibrin accumulation may be implicated in the genesis of pulmonary edema after embolization in rats, although undoubtedly fibrin accumulation and its degradation products play a key role since the extra- vascular lung water content in the heparinized rats was reduced by approxi- mately 50% of the value in the non-heparinized rats. In summary, heparin pretreatment is not as effective in preventing the extravascular lung water accumulation after pulmonary microembolism in rats as is in dogs. This

difference may be due to the greater release and activation of other humoral factors in the rat lung occurring independent of fibrin accumulation.

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