Disseminated Intravascular Coagulation inDiabetes Mellitus, with Reference to the Role

of Increased Platelet AggregationHau C. Kwaan, M.D., John A. Colwell, M.D., and Nibha Suwanwela, M.D., Chicago

SUMMARY

A state of hypercoagulability associated with changes inthe clotting factors and platelet function has been postulatedto be important in the increased thrombotic complicationsin diabetes mellitus. A fifty-nine-year-old man was admittedwith severe diabetic ketoacidosis and shock. Shortly afterrestoration of his gross fluid and electrolyte imbalance, hedeveloped extensive bleeding into his lungs and gastroin-testinal tract. His serum was grossly lactescent. His bloodwas incoagulable. The existence of disseminated intravas-cular coagulation was indicated by a decrease in the plateletcount and in the levels of fibrinogen, plasminogen, andFactors V and VIII. Plasminogen activator, plasmin, andfibrinogen degradation products all were increased in theplasma, as were cholesterol, triglycerides, phospholipids,total lipids, and free fatty acid concentrations. Postmortemexamination revealed widespread hemorrhages. Micro-thrombi characteristic of platelet aggregates were present inhis lungs and kidneys. Serial dilutions of his lipidemicplasma up to 1:150 resulted in increased ADP-induced ag-gregation of normal platelets. We postulate that the stronginfluence of the patient's plasma on platelet aggregation wasthe initial event leading to his disseminated intravascularcoagulation. The finding of a platelet aggregation-enhancingfactor in other diabetic subjects may shed light on themechanism of thrombogenesis in diabetes. DIABETES 21:108-13, February, 1972.

Recent reports on studies of blood coagulation factorsand platelet functions in diabetes mellitus indicate thata hypercoagulable state may exist in these subjects, par-

From the Department of Medicine, Northwestern UniversityMedical School, and the Hematology and Metabolism Sections,Veterans Administration Research Hospital, Chicago, Illinois60611.

Address requests for reprints to: Hau C. Kwaan, M.D.,Hematology Section, Veterans Administration Research Hos-pital, 333 East Huron Street, Chicago, Illinois 60611.

ticularly in those with ischemic heart disease.1"4 Thehypercoagulability is associated with shortened plasmacephalin time, increase in Factors V and VIII, andfibrinogen levels,1>3 and increased platelet adhesiveness.3

Increase in Platelet Factor 4 has also been found duringplatelet aggregation in diabetic patients.5 Such changesmay play an important role in the thrombotic compli-cations in diabetes. However, instances of extensivethrombosis, such as those seen in acute disseminatedintravascular coagulation, have not yet been reported indiabetic subjects. The present paper describes the clini-cal and postmortem findings of a patient with diabetesmellitus, who died from disseminated intravascular coag-ulation and whose plasma was found to greatly enhancethe ADP-induced aggregation of normal platelets.

CASE REPORT

H. D., a fifty-nine-year-old Negro male, was diagnosed ashaving diabetes mellitus in 1966. He was treated with tolbuta-mide, 500 mg. daily. Three years later he was seen in theemergency room of the Veterans Administration ResearchHospital in a semicomatose state with a history of abdominalpain and vomiting for three days. Physical examination re-vealed that the patient responded only to painful stimuli, hada regular pulse rate of 112/min., blood pressure of 60/40 mm.Hg, respiratory rate of 44/min., and a temperature (rectal)of 1050 F. He was dehydrated and his breath smelled of ace-tone. Nuchal rigidity was present. His chest and abdomen werenormal. Tendon reflexes in the lower limbs were absent. Initiallaboratory findings were: hematocrit 48.5 per cent, hemoglobin18.8 gm. per cent, white cell count 5,000/mm.3. The peripheralblood smear showed normal red cell morphology, with ade-quate numbers of platelets. Blood sugar was 860 mg./ioo ml.,blood urea nitrogen 60 mg. per cent, serum sodium 150mEq./L., serum potassium 3.0 mEq./L., chloride 114 mEq./L.,and CO.2 11.6 mEq./L. His serum acetone was strongly posi-tive in a 1:1 dilution and the serum was found to be lactescent.In spite of a possible increased viscosity of the blood, no diffi-culty was encountered when blood was withdrawn with a 20-Gneedle. His urine was strongly positive for sugar and acetone.A lumbar puncture was performed, and a cerebrospinal fluidwas found to have a pressure of 200 mm. H2O. It was clear,

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colorless and contained no cells. A chest roentgenogram wasnormal. Cultures of his cerebrospinal fluid, blood, and urinewere negative.

The patient was given vigorous replacement therapy toachieve a state of fluid and electrolyte balance. In the firstnine hours, he received 6 L. of 0.45 per cent saline solution,2 L. of 5 per cent dextrose solution, 120 mEq. of sodium bicar-bonate, 20 mEq. of potassium chloride, and 325 U. of Regularinsulin (one half by the intravenous route, and one-half sub-cutaneously). His urine output was 250 ml. during this period.The systolic blood pressure was maintained at 90-100 mm.Hg by Levarterenol. His blood sugar fell to 250 mg./ioo ml.and CO2 rose to 18 mEq./L. nine hours after admission. Hishematocrit fell to 42.5 per cent while hemoglobin remained at19 gm. per cent. The latter value was attributed to themarked lactescence of his blood and should be regarded as afalsely high figure. His white cell count was 11,000/mm.3 Hebecame more alert and responded to verbal command. How-ever, his blood obtained at this time did not clot. Specialstudies of his clotting factors showed a platelet count of10,000/mm.3, prothrombin time of over 120 sec, partialthromboplastin time of over 120 sec, Factor VIII6 level of8 per cent, Factor V7 level of less than 10 per cent, thrombintime8 of over five minutes, fibrinogen9 level of 165 mg. percent, plasminogen10 level of 1.7 CTA (Committee on Throm-bolytic Agents, National Heart Institute) U./ml. (normalvalue 7.8 ± 1.6), plasminogen activator of 3.1 CTA U./ml.11

(normal value 0-1.0 CTA U./ml.), and plasmin level of 0.8CTA U./ml.10 (normal value unmeasurable by this technic).His serum was still strongly lactescent. Serum cholesterolwas 1,550 mg. per cent, triglyceride 15,780 mg. per cent,phospholipid 1,645 mS. per cent, and total lipids 19,690 mg.per cent. The free fatty acids done by Trout's modification12

of Dole's method13 was 1.689 mEq./L. (normal range 0.3-0.7 mEq./L.). Due to the massive chylomicronemia, definite

delineation of beta and pre-beta lipoproteins on electrophoresiscould not be obtained on his plasma. However, when thiswas done on diluted plasma, a predominance of origin mate-rial was found and the pattern was most compatible with TypeI. He soon began to bleed from his gums, nose, and intosubcutaneous tissues at various parts of the body. His urineoutput dropped to a total of 50 ml. during the second ninehours after admission. His condition deteriorated, and heexpired eighteen hours after admission. Postmortem examina-tion revealed widespread hemorrhages in the liver, lungs, kid-neys, adrenals and the gastrointestinal tract. The blood in thelarge vessels was fluid and not clottable. Microscopic exami-nations of his viscera revealed that microthrombi were num-erous in the lungs and kidneys (figures 1 and 2). Thesethrombi had the histochemical characteristics of platelet aggre-gates, being stained intensely pink with periodic-acid-Schiff(PAS) technic and very lightly blue with phosphotungstic-acid-hematoxylin.Platelet aggregation studies

Serial dilutions of the patient's citrated plasma obtainedimmediately before death were made with normal saline.0.1 ml. of these samples were mixed with 0.8 ml. normalplatelet-rich plasma with platelet counts of 400,000/mm.3

and the mixtures were studied for platelet aggregation bythe addition of 0.1 ml. adenosine diphosphate (final con-centration in the mixture was 1.05 x io '6 M). 0.1 ml. salineinstead of patient's plasma was also used to mix with thenormal platelet-rich plasma to serve as control. The aggrega-tion was carried out in a Chronolog Model S201 Aggrego-meter and the pattern recorded by a Bausch and Lomb 10 mvrecorder (Model VOM 5). A marked increase in the secondphase aggregation over the control was noted in the samplescontaining patient's plasma up to a final concentration of1:150. Observations on the increase of the aggregation in thedifferent dilutions over the control four minutes after addition

FIGURE I

Microthrombi present in patient'slung. Hematoxylin and eosin stain.

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DISSEMINATED INTRA VASCULAR COAGULATION IN DIABETES MELLITUS

FIGURE 2

Microthrombus present in the juxta-glomerular arteriole in patient's kid-ney. Hematoxylin and eosin stain.

of ADP are shown in figure 3 and table 1. In the sampleswith patient's plasma more concentrated than 1:50, the ag-gregation pattern could not be measured since the markedlactescence interfered with the optical transmittance in theaggregometer.

Another plasma sample drawn four hours before patient's

minutesFIG. 3. The effect of patient's plasma on the ADP-induced ag-

gregation of normal platelets. The upper aggregationpattern was performed on a mixture of O.I ml. of 1:50dilution of patient's plasma with 0.8 ml. of normalplatelet-rich plasma. The lower aggregation pattern isthat of control mixture; normal saline replaced patient'splasma.

death was diluted 1:50 and tested similarly for its effect onthe ADP-induced aggregation of normal platelets. It wasfound that an increase of 45 per cent of the aggregation overthe control occurred at four minutes after the addition ofADP.

The patient's plasma diluted 1:50 when mixed with normalplatelet-rich plasma 1:9 v/v did not cause spontaneous aggre-gation without the addition of ADP.

TABLE 1The effect of different dilutions of patient's plasma on the

ADP-induced aggregation of normal platelets*

Final cone, in normalplatelet-rich plasma

% aggregationincrease (AT.)

0:1(control)

0

1:50

43

1:80 1:150

29 17

* The enhancement of the aggregation of normal plateletis expressed as the increase in percentage aggregation overthe controls at four minutes after the addition of ADP(AT4, see figure 3). Such enhancement is directly propor-tional to the strength of the patient's plasma.

DISCUSSION

Disseminated intravascular coagulation (DIC) isknown to occur in a wide variety of conditions. Amongthose that have a possible causal relationship to thepresent case are septicemia,14 acidosis,15 and shock.16

The possibility of septicemia is unlikely since repeatedcultures of patient's blood, urine, and cerebrospinal fluid

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DIABETES MELLITUSINSULIN LACK

KETOACIDOSIS

SHOCK

DECREASEDLIPOPROTEIN LIPASE

INCREASEDNOREPINEPHRINE

INCREASED^-LIPOPROTEIN

\

PLATELET AGGREGATION] ACTIVATION OFFACTORS XI , XII

FIGURE 4

A diagrammatic reconstruction of theprobable chain-of-events leading todisseminated intravascular coagulationin the patient.

DISSEMINATED INTRAVASCULAR COAGULATION

were negative, and no evidence of infection was foundon postmortem examination.

Judging by venous CO2 values and qualitative plasmaketone determinations, the patient had severe keto-acidosis. DIC appeared at a time when most of themetabolic abnormalities had been corrected with fluidsand insulin. Thus, it is unlikely that acidosis could haveplayed a significant part in precipitating DIC. The sameis true with hemoconcentration and hyperglycemia whichhe had on admission. However, the patient was main-tained on norepinephrine throughout the hospital course.This continuous stimulation of the a-adrenergic receptorsites could also have contributed to the development ofDIC as had been shown by Whitaker et al. in experi-mental catecholamine shock.17

At the onset of clinically evident DIC in this patient,the analysis of the lactescent plasma revealed substan-tially raised levels of cholesterol, triglycerides, phos-pholipids, total lipids and free fatty acids. Since theinstitution of insulin therapy in diabetic ketoacidosiswould produce a rapid fall in the free fatty acid level,18

it is possible that this patient's free fatty acid level waseven higher on admission to the hospital. On the otherhand, norepinephrine infusion tends to raise the plasmafree fatty acid level.19 Futhermore, free fatty acid couldbe mobilized by injection of a large dosage of ACTHin experimental animals and resulted in shortened clot-ting time and widespread thrombosis.20 Since plasmacortisol could be markedly elevated due to impairedmetabolism,21 our patient's free fatty acid concentrationwould remain high throughout the course of his ill-

ness. These high levels undoubtedly contributed to hisDIC. The thrombogenic effect of fatty acids in vivo andthe clot promoting properties in vitro have been exten-sively studied.22"27 Intravenous infusions of long-chainsaturated fatty acids in experimental animals have re-sulted in widespread intravascular thrombosis and death.Upon histological and electron microscopic examinationit has been shown that the thrombi resembled naturalthrombi rather than blood clots and contained large ag-gregates of platelets.25 In vitro studies have demon-strated that long-chain free fatty acids can accelerateclotting by activation of Factor XII (Hageman Factor)and of Factor XI.22-28-29 These fatty acids also havedirect aggregating action on platelets.30-31 In addition,they were also found to" enhance the platelet aggrega-tion induced by ADP and thrombin.30"34 The enhancedaggregation was found to be related to the saturationof free fatty acids and the free fatty acid:albumin ratio.32

Short-chain saturated fatty acids and unsaturated fattyacids have no effect. Haslam30 suggested that the plate-let aggregation effect was mediated by ADP releasedfrom the platelets by the fatty acids. Thus, we suspectthat elevated plasma free fatty acid levels contributedto platelet aggregation in this patient.

An influence of other lipids on platelet aggregationhas also been described.35 Among the different lipopro-tein fractions, beta-lipoprotein has the strongest en-hancing effect on platelet adhesiveness and aggregation.Chylomicra will also increase platelet adhesiveness, butonly at high concentrations. Although lipoproteins werenot quantitated in the present study, the marked eleva-

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tions in plasma levels of cholesterol and phospholipidssuggest some elevation of beta-lipoproteins. However,lipoprotein electrophoresis of diluted plasma showed amarked excess of origin material (chylomicra), andplasma lipid analysis showed unusually high levels oftriglycerides. This suggests that in our patient, chy-lomicra may have contributed greatly to increased plate-let aggregation.

Another factor which might contribute to DIC ishyperglycemia. D-glucose could increase platelet adhe-siveness and was believed to be responsible for the in-creased platelet stickiness in diabetes.36 Patient's initialblood sugar of 860 mg./ioo ml. could indeed result inhis platelet stickiness.

We thus conclude that platelet aggregation in ourpatient was of prime importance in precipitating theonset of his DIC. This conclusion was supported by thepostmortem findings in his lungs and kidneys. Whileour studies do not allow a precise description of thechain of events, a postulated sequence leading to theDIC is shown in figure 4.

The clinical events leading to platelet aggregation andthe DIC syndrome were profound in the subject studied.It seems likely that further investigation into plateletaggregation in subjects with less severe diabetes mellitusmight be productive. In particular, correlation of serumlipids with platelet aggregation are warranted. Prelimi-nary studies37-38 to be reported in detail elsewhere sug-gest that a platelet aggregation-enhancing factor may befound in the plasma of many diabetic patients. The sig-nificance of such increased platelet aggregation inthrombogenesis in diabetes remains to be determined.

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