The Effect of pH on Platelet and Coagulation Factor Activities

Chaim Chalmoff, MD, Petah-Tiqva, Israel Draga Creter, MSc, Petah-Tiqva, Israel Meir Djaldetti, MD,’ Petah-Tiqva, Israel

The successful

control of gastrointestinal bleeding is important because of its associated high mortality in patients with gastric, duodenal, or stress ulcers [1,2]. Appropriate conservative therapy includes cooling [3], vasoconstriction [4,5], or change in the pH of the intragastric content by gastric lavage with alkaline solutions. .tn the successful experience of Curtis et al [2], the alkaline solution used was magnesium aluminum hydroxide. We tried the same method with a slight change, using an alkaline solution of 5 liters of cold water with sodium bicarbonate of an amount sufficient to keep the pH of the solution between 7.5 and 8.0 and the pH of the escaped water after the lavage between 7 and 7.5. The exact mechanism by which alkalinization of the gastric content decreases local bleeding tendency is not clear. It may be postulated that the increased pH induces changes in the platelet functions or affects also the clotting mechanism at the bleeding site. The present paper studies the influence of the base pH on the coagulation mechanism. Material and Methods Ten healthy volunteers were used for pH blood studies. Venous blood, 18 ml, was anticoagulated with 3.8 per cent sodium citrate, 2 ml, and the platelets were counted. Platelet-rich plasma was obtained by centrifugation at 85 X g for 10 minutes and the pH determined. The plateletrich plasma was divided in equal portions in plastic tubes, and in each tube the pH was adjusted with tenth-normal hydrochloric acid to 5.0,6.0,6.5, and 7.0, and with sodium bicarbonate to 8.0. The initial pH of the platelet-rich From the Departments of Surgery “A” and Internal Medicine “8”. and Clinical Laboratories, Hasharon Hospital, Petah-Tiqva, Tel Aviv University Medical School. Tel Aviv. Israel. Repiint reque& should be addressed to Meir Djaidetti, MD, Department of internal Medicine “B”, Hasharon Hospital, Petah-Tlqva, Israel. ’ Estabilshed investigator of the Chief Scientist’s Bureau. Ministry of Health. Israel.

Volume 136, August 1978

plasma was found to range between 7.5 and 7.6. The remaining plasma was again centrifuged at 800 X g for 10 minutes, and the platelet-poor plasma obtained was used for the coagulation studies. The platelet count was carried out with an improved Neubauer chamber using a light microscope [S]. The platelet aggregation pattern was recorded with an Evans Electroselenium Platelet Aggregation Meter Model 169, with recorder, using 0.6 ml platelet-rich plasma and 6 X 10e2 PM epinephrine and a minimal recording time of 6 minutes [ 71. Platelet-rich plasma was considered as 0 per cent and platelet-poor plasma as 100 per cent. The maximum percentage of aggregation was calculated as well as the normal double wave pattern appearance. The platelet calcium determination was carried out by an atomic absorption spectrophotometry method [8] and platelet serotonin by a fluorescence method [9]. Platelet factor III availability was assayed by the method of Hardisty and Hutton [IO]. Prothrombin time was determined by a one stage method [II]. Kaolin-activated partial thromboplastin time (Hyland partial thromboplastin) [12] and fibrinogen were determined by an immunologic method, using self-made plates with Behringwerke’s fibrinogen antiserum [13]. All pH determinations were done on a Radiometer A/S-Acid Base Cart Type ABC1 (Copenhagen NV, Denmark) [14].

Results

The results of platelet calcium determination, platelet serotonin content, and platelet factor III availability are shown in Table I. Platelet calcium release increased directly with the pH, until pH 7.0 was reached. At pH 7.6 and 8.0 there was a decrease, the amount of calcium being similar to that found at pH 6.5. Platelet serotonin showed maximum activity at pH 6.5, whereas at pH 7.0,7.6, and 8.0 there were no essential differences, the values being at the upper limit of normal. Platelet factor III availability was normal only at pH 7.0 and 7.6.

257

Chaimoff, Creter, and Djaldetti

TABLE I

Determlnatlon of Platelet Functions at Different pH Levels

pf-f

Levels in Platelets Calcium Serotonin (ng/200,000) (Mgll miliard)

5.0 6.0 6.5 7.0 7.6 8.0

6.55 9.10 9.63 11.43 10.64 10.10

Normal range

f f f f f f

0.24 0.19 0.17 0.18 0.54 0.24

9-15

0.40 0.42 0.69 0.56 0.63 0.55

f f f f f f

0.031 0.011 0.018 0.017 0.005 0.025

0.13-0.77

Platelet Factor Ill Availability (set) >I80 107 f 2.58 49.5 f 0.45 39.1 f 0.41 42.6 f 0.34 62.3 f 0.92 35-45

8.0

1.0

1.6

Note: All values are given as the mean f standard error of the mean.

TABLE II

1.4

Coagulatlon Parameters at Different pH Levels Partial Thromboplastin Time

pf-f

Prothrombin Time (set)

5.0 6.0 6.5 7.0 7.6 8.0

>60 31.9 f 1.86 25.2 f 0.17 16.8 f 0.10 13.7 f 0.10 17.4 f 0.21

>120 69.6 f 0.64 51.9 f 0.56 46.4 f 0.36 39.0 f 0.34 49.6 f 0.77

Normal ranae

13-15

35-45

(set)

Fibrinogen (mg/df) 270 348 348 322 308 270

f f f f f f

2.96 2.83 2.83 3.32 3.38 2.96

200-400

Note: All values are given as the mean f standard error of the mean.

The coagulation parameters at different pH levels are given in Table II. The prothrombin time was in the normal range only at pH 7.6 and at the lowest activity at pH 6.0, whereas at pH 5.0 it was not detectable. The same results were found for the partial thromboplastin time. The fibrinogen values were within the normal range at each pH determination. Figure 1 represents the platelet aggregation patterns induced by epinephrine at different pH levels. A normal double wave pattern was found only at pH 7.6 and 7.8. In all determinations the platelet count was 211,300 f 2,780 (SE). Comments

Stress ulcer and hemorrhagicgastritisare the most severe forms of upper gastrointestinal bleeding and the most difficult to control. LeVeen et al [I] emphasized their high mortality (more than 50 per cent). In addition to the various procedures proposed

250

1.0

6.5

Ffgure 1. Epinephrine-induced platelet aggregation at different pH /eve/s.

for the management of upper gastrointestinal hemorrhage, Curtis et al [2] suggested control of the intragastricpH at 7.0. They achieved arrest of massive upper gastrointestinal bleeding in twenty-three of twenty-five patients by applying this procedure. In one patient in whom the pH could not be increased to more than 4.5 and who therefore did not respond to treatment, the bleeding ceased only after operation. In patients in whom the hemorrhage stopped at pH 7.0, it reappeared when the pH decreased to 5.0. The bleeding subsided with additional increase of the intragastric pH to 7.0. One possible explanation of the beneficial effect of the elevated pH on bleeding control forwarded by the authors was decreased back-diffusion of hydrogen ions through the

The Amerlcan Journal of Surgery

pH and Platelet Coagulation Factors

gastric mucosa, or total and complete inhibition of pepsin acbivi ty. The present study shows that alteration of the pH aifects some of the platelet functions as well as the prothrombin and partial thromboplastin times. McLean and Veloso [ 251have shown that at pH 5.8 rabbit platelets acquire a more symmetrical form and the optical density of the platelet suspension increases. With an increase in the pH, platelet. aggregation rapidly increases. The authors concluded that a lower pH level prevents aggregation, either by a change of the platelet shape or by prevention of the activation of plasma factors required for aggregation. Dependence of platelet aggregation upon pH was demonstrated also by Rogers [16]. In his experiments plasma acidosis caused inhibition of epinephrineinduced platelet aggregation, whereas alkalosis enhanced this platelet function. Our results support this observation. In addition, other platelet functions changed with variations of the pH. The platelet calcium and serotonin content increased markedly with elevation of the pH, reaching a maximum at pH 7.0 and 6.5, respectively. We do not have an explanation for this phenomenon, except that membranal or intracellular changes induced by a lower pH may account for the difference in calcium and serotonin release. It is of interest that platelet factor III showed marked changes even with small variations of the pH. The prothrombin time was markedly prolonged at lower pH levels, but it became shorter with correction of the pH to normal. Alkalosis did not alter significantly the prothrombin time. A similar pattern was observed with partial thromboplastin time. A possible explanation for these results could be a change in the fibrinogen level at different pH levels, but the lack of significant variations in the fibrinogen level at low and elevated pH observed in the present work does not support this assumption. We conclude that elevation of the pH may induce an activation of certain platelet functions and some plasma factors. It is possible that this is one of the mechanisms by which lavage of the stomach with alkaline solutions controls intragastric hemorrh,age. Summary

After observing that lavage of the stomach with alkaline solutions has a beneficial effect on the control of gastric hemorrhage, we examined the platelet

Volume 136, August 1979

and coagulation factor activities under conditions of lower and elevated pH. The results showed that change of the pH from acidosis to 7.0 and even to slight alkalosis induces platelet aggregation, platelet calcium and serotonin release, as well as platelet factor III availability. The prothrombin and partial thromboplastin times approached normal levels, whereas the fibrinogen level did not change significantly. The results obtained may serve as an explanation for the control of the intragastric bleeding in patients treated by maintenance of t,heir gastric pH at. 7.0. References 1. LeVeen HH, Falk G, Diaz C, Wynkoop B-J, Piccone VA, Yarnoz

2.

3.

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6. 7. 8.

9. 10.

11. 12.

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14.

15.

16.

MD, Langsam AA, Nelson JH, Pedowitz WJ, Belfasky RB: Control of gastrointestinal bleeding. Am J Surg 123: 154, 1972. Curtis LE. Simonian S, Buerk CA, Hirsch EF, Soroff HS: Evaluation of the effectiveness of controlled pH in management of massive upper gastrointestinal bleeding. Am J Surg 125: 474,1973. Sosin H. Bernstein EF, Madsen A, Fusino K, Wangensteen OH: Mechanism of control of hemorrhage from duodenal ulcer by gastric hypothermia. JAMA 186: 219. 1963. Nusbaum M, Baum J, Blakemore WS: Clinical experience with the diagnosis and management of gastrointestinal hemorrhage by selective mesenteric catheterization. Ann Surg 170: 506, 1969. Rosch J, Gray RK, Grollman JH, Ross G, Steckel RJ. Weiner M: Selective arterial drug infusions in the treatment of acute gastrointestinal bleeding. Gasfroenferology 59: 341, 1970. Dacie JV. Lewis SM: Practical hematology, 5th edition. Edinburgh, Churchill Livingstone, 1975. Born GVR, Cross MJ: The aggregation of blood platelets. J Physiol168: 178, 1963. Creter D. Rubinstein J, Menache R: A simple and rapid platelet calcium determination. Normal values and results in MayHegglin anomaly. Acta Haematol57: 168, 1977. Drummond AH, Gordon JL: Rapid, sensitive microassay for platelet 5HT. Thrombos Diathes Haemorrh 31: 366, 1974. Hardisty RM, Hutton RA: The kaolin clottina time of olatelet-rich plasma: a test of platelet factor-3 availability. B;J~Haemato/ 11: 258. 1965. Quick AJ: The hemorrhagic diseases and the physiology of hemostasis. Springfield, Charles C Thomas. 1942. Proctor RR, RapapoiSI: The partial thromboplastin time with kaolin. A simple screening test for first stage plasma clotting factor deficiencies. Am J C/in P&ho/ 36: 212, 1961. Mancini G, Carbonara AO, Heremans JF: lmmunochemical quantitation of antigen by single radial immunodiffusion. Immunochemistry 2: 235, 1965. Winters RW, Engel K, Dell RB: Acid Base Physiology in Medicine. A Self-Instruction Program, 2nd edition. Ohio and Copenhagen, London Company and Radiometer A/S, 1969. McLean JR, Veloso H: Change of shape without aggregation caused by ADP in rabbit platelets at low pH. Life Sci6: 1983, 1967. Rogers AB: The effect of pH on human platelet aggregation induced by epinephrine and ADP. Proc Sot Exp Viol Med 139: a100, 1972.

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The effect of pH on platelet and coagulation factor activities.

The Effect of pH on Platelet and Coagulation Factor Activities Chaim Chalmoff, MD, Petah-Tiqva, Israel Draga Creter, MSc, Petah-Tiqva, Israel Meir Dj...
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