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ORIGINAL ARTICLE
Year : 2012  |  Volume : 6  |  Issue : 1  |  Page : 35-40

Glanzmann's thrombasthenia: The clinicopathological spectrum and its diversities


1 Department of Pathology, Kasturba Medical College, Manipal, India
2 Department of Pathology, Melaka Manipal Medical College, Manipal, Karnataka, India

Date of Web Publication28-Aug-2012

Correspondence Address:
Sushma Belurkar
Department of Pathology, Kasturba Medical College, Manipal, Karnataka
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0331-3131.100220

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   Abstract 

Aim: To study the clinical spectrum of patients with Glanzmann's Thrombasthenia (GT) in a tertiary care hospital in South India and to correlate the clinical & laboratory parameters of GT.
Materials and Methods: This study includes 15 patients who were diagnosed as GT by characteristic clinical features, basic lab parameters (prolonged bleeding time, abnormal clot retraction and absence of platelet clumps on direct smear) and platelet aggregometry.
Results: 66.6% of our patients were below 10 years of age. The common forms of bleeding in our patients was epistaxis,gum bleeding and prolonged bleeding from trauma site which was seen in >50% of our patients.The significant findings on routine laboratory testing was a prolonged bleeding time (93.33%) and an abnormal clot retraction test (66.6%). Platelet aggregation studies showed decreased aggregation with ADP, Collagen and Epinephrine in 100% of our patients whereas 50% also showed decreased aggregation with Ristocetin.Hence most of the patients in this study presented with the classical symptoms and laboratory findings of GT however some had unusual findings.
Conclusion: The prevelance of GT is higher in southern India due to increased incidence of consanguineous marriages in this area.Though basic hemostatic screening tests and platelet aggregometry are sufficient to establish the diagnosis in majority of the patients,however some may require more sophisticated tests like flowcytometry for glycoprotein (GP) IIb-IIIa.

Keywords: Glanzmann′s thrombasthenia, Platelet aggregation, Glycoprotein IIb-IIIa, haemostasis, mucocutaneous bleeding, consanguinity


How to cite this article:
Belurkar S, Kurien A, Singh PB. Glanzmann's thrombasthenia: The clinicopathological spectrum and its diversities. Ann Nigerian Med 2012;6:35-40

How to cite this URL:
Belurkar S, Kurien A, Singh PB. Glanzmann's thrombasthenia: The clinicopathological spectrum and its diversities. Ann Nigerian Med [serial online] 2012 [cited 2020 Aug 10];6:35-40. Available from: http://www.anmjournal.com/text.asp?2012/6/1/35/100220


   Introduction Top


Glanzmann's thrombasthenia (GT) was first described by a German pediatrician Dr. Glanzmann in 1918. He designated it as "hereditary hemorrhagic thrombasthenia." [1] This disorder is inherited in a classic Mendelian autosomal recessive fashion and is the most common inherited abnormality of platelets. Its prevalence is high in areas with high consanguinity. [2]

The concept of GT was first proposed in 1970s when Nurden and Caem demonstrated that platelets from patients with GT had selective abnormalities in their membrane glycoprotein complex (1). This led to recognition that the disease was provoked by specific deficiency of GP IIb-IIIa. It was later established that this complex was a member of ubiquitous integrin family of cell surface receptors. [1] The genes for GP IIb-IIIa are located on the long arm of chromosome 17(17;q21-22).

GT has a very uneven geographic distribution which is demonstrated in the review article by George

et al., [3] which states that 12 of 177 patients were from the United States, 55 were from Israel and Jordan, and 42 were from South India. A study done in Iran has reported a prevalence rate of 1:200,000 among a population of 60 millions. [4]

GT has been classified into three types: [5]

Type I: GP IIb-IIIa levels <5%, no platelet aggregation, no clot retration, no a granule storage pool of fibrinogen.

Type II: GP IIb-IIIa levels 5-15%, no platelet aggregation, residual clot retraction, readily detectable but often subnormal a granule pools of fibrinogen.

Type III: Normal levels of dysfunctional GP IIb-IIIa, no or very abnormal platelet aggregation, fibrinogen storage, and clot retraction variably affected.

GT patients usually present with mucocutaneous bleeding such as purpura (86%), epistaxis (73%), menorrhagia (98%), or gingival bleeding (55%). [3] However, postsurgical bleed and gastrointestinal bleed can be serious. [2] It is marked by prolonged bleeding time, normal platelet count, abnormal clot retraction, and absent platelet clumps on direct smear. Platelet aggregation response is lacking to all platelet agonists except ristocetin. [6]

Although platelet membrane GP defects and genetic defects were well documented, a wide heterogenecity is recognized in these disorders. The main objective of this study was to evaluate the diverse spectrum of this disorder and to study the magnitude of this disease in a tertiary care center in South India.


   Materials and Methods Top


A retrospective study of 15 patients diagnosed with GT in the Haematology Laboratory of Kasturba Medical College, Manipal, from June 2006 to June 2008 (2 years) was undertaken. The clinical details and the various laboratory investigations done were reviewed and analyzed.

In the clinical data, the points that were stressed upon were age of the patient, sibling and family history, consanguineous marriage between parents, sites of bleeding, and the severity of bleeding.

The laboratory parameters studied included hemostatic screening tests and platelet aggregation studies with ADP, collagen, epinephrine, and ristocetin. Von Willebrand factor assay was also done in only 11 patients due to financial constraints.

The hemostatic screening tests done were:

  1. Bleeding time (BT) by modified Ivy's method (N 1-6 minutes)
  2. Prothrombin time (PT) and activated partial thromboplastin time (APTT) (N PT 12-16 sec and APTT is 30-35 sec)
  3. Thrombin time (TT) (N 15-22 sec)
  4. Clot retraction test by the conventional method (N is >50%)
  5. Urea solubility test for factor XIII
  6. Platelet count by automation (Sysmex KC4 or Pentra 80 XL) (N range is 140-400 × 10 3 /cmm)
  7. Direct finger prick smear to look for platelet clumps
Blood samples for these tests were collected in 3.8% Sodium Citrate vacutainer. EDTA sample was also collected for platelet count and direct smear was prepared from finger prick. The coagulation tests were performed on Sigma Amelung Semi Automatic Analyser by mechanical method for end point detection. Routine precautions required for coagulation tests were taken.

Platelet aggregation tests were done on Chronolog Aggregometer 490 with reconstituted ADP

(10 μM/ml), collagen (2 μg/ml), epinephrine (10 μM), and ristocetin (1 mg/ml). Four tubes of blood were collected in 3.8% Sodium Citrate (4.5 ml each) and one tube with EDTA for platelet count. The blood samples were processed within 2 hours of collection. 10 ml of blood was centrifuged at 1000-1200 rpm for 7-8 minutes to obtain platelet-rich plasma (PRP) with a platelet count of 2-3 lakhs and the remaining sample was centrifuged at 3500 rpm to obtain platelet poor plasma (PPP).

Samples from normal patients were treated similarly to prepare PRP and PPP and ran simultaneously to serve as controls. Baseline optical density was set at 0 with PRP and at 100% with PPP. The percentage of aggregation was compared with controls and the normal reference ranges provided by the company.

Von Willebrand assay was done by ELISA method as per standard techniques.


   Results Top


The age of the patients in this study ranged from 6 months to 22 years. Out of the 15 patients studied, 10 (66.6%) were below the age of 10 years.

Four patients (26.6%) had history of consanguinity in parents and 2 had positive sibling history. Male:female ratio was 1:1.

86.6% of our patients presented with mucocutaneous bleeding. The commonest form of bleeding was epistaxis and gum bleeding followed by easy bruisability and prolonged bleeding from trauma site. Four females in this study were in the reproductive age group and all of them presented with menorrhagia, hence menorrhagia could be a more common form of bleeding in females who have reached menarche. Only two patients had more serious forms of bleeding [Table 1].
Table 1: Spectrum of clinical presentation in the patients (n = 15)

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All 15 patients had a normal platelet count, normal PT, APTT, TT, and a normal factor XIII test as was expected in GT.14 patients had a prolonged BT and 10 patients had an abnormal clot retraction test, whereas 1 patient showed a normal BT and 5 patients had normal clot retraction. Eleven patients had absence of platelet clumps on direct smear but four had small clumps [Table 2].
Table 2: Hemostatic screening tests in the patients (n = 15)

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Platelet aggregation study results were available only for 13 patients and all of them showed decreased aggregation with ADP, collagen, and epinephrine [Figure 1].
Figure 1: Platelet aggregation pattern seen in GT patients with ADP,Collagen and Epinephrine Patient (a) and control (b)

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Seven patients showed normal aggregation with ristocetin, but six patients (53.33%) showed decreased aggregation with ristocetin also [Figure 2] and [Figure 3].
Figure 2: Platelet aggregation pattern in GT showing normal aggregation with Ristocetin but decreased aggregation with other agonists in patient (a). Control (b)

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Figure 3: Platelet aggregation pattern showing normal aggregation with Ristocetin in control (a) and disaggregation pattern in patient (b)

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Von Willebrand assay was done in 11 patients and was within normal limits for all 11 patients.


   Discussion Top


Our study basically reflects the diverse clinical features and laboratory findings in GT. The prevalence of GT is higher in South India when compared with other parts of India due to the increased incidence of consanguineous marriages. The largest study done in India is from CMC, Vellore, which has reported 42 cases of GT in a 14-year period from June 1966 to June 1980 and they have reported consanguinity in 71% of their cases. [7] A study done in Western India which includes 120 women in the reproductive age group with menorrhagia has reported GT in only 3.3% of their cases. [8] The incidence of consanguineous marriages is 21% in this region of India. [9]

The age at diagnosis is varied, but most cases are diagnosed at an early age. Various studies done in India and abroad have shown that GT is a disease of children and young adults with majority of patients being less than 20 years. [2],[3],[6] The age of patients in our study ranged from 6 months to 22 years. Sixty six percent of our patients were below 10 years of age and 93.33 % were below 20 years. The male to female ratio is 1:1 in this study just like studies done in India and other parts of the world. [3],[6] However, Badhe et al. in their study have shown markedly high incidence of GT in females. [2]

Most patients in our study presented with mucocutaneous bleeding like epistaxis, gum bleeding and skin bleeds. Menorrhagia was seen in four female patients. Finding is supported by other studies as well. [2],[3],[6] Joint bleeds as seen in coagulation disorders is rare in these patients.

Consanguinity between parents has been found to be a very important risk factor for GT. Hence, the incidence is high in ethnic groups like Iraqi Jews, [10] Northern Iranis, [11] and Southern Indians. [7] Our study showed consanguinity in 26.6% of patients. However, there is a wide variability in the consanguinity pattern in the different studies. A study done in Iran showed an incidence of consanguinity as high as 93.3% [4] in their GT patients, whereas in Pakistan the incidence was only 14%. [6]

The important screening tests which help in diagnosis of GT are prolonged bleeding time, absent/decreased clot retraction, and absence of platelet clumps on direct smear. The present study showed prolonged bleeding time in 93.33% of patients and abnormal clot retraction in 66.66% of patients. This is well supported by other studies. Chediak et al. and Badhe et al. have shown bleeding time more than 10 minutes and abnormal clot retraction in 100% of their patients. [2],[10]

Prolonged BT is an indication of delay in formation of primary hemostatic plug due to defective platelet aggregation. Although BT is an easily available tool for assessing in vivo platelet function but its high operator dependency and lack of precision are the major obstacles in providing meaningful results. [6] Use of PFA-100 system can replace BT tests for GT. [1] The PFA measures the closure time when blood is passed through collagen-based filters under high shear stress, blood from GT patients fail to plug the filters, resulting in prolonged closure time.

Activated platelets interact with fibrin within the clot and contraction of platelet cytoskeleton generates force for clot retraction. [12] This explains absent or reduced clot retraction in GT patients. Prolonged BT and reduced or absent clot retraction in GT has been a consistent finding in other studies as well. [2],[13]

One patient in this study had a normal BT. This was initially thought to be a technical error in the performance of BT which is well known. It was interesting to note that in this patient the clot retraction was also found to be normal. Hence, later it was thought that this patient could be the variant form of GT. Besides this patient, four other patients had a normal clot retraction test but an increased BT. These patients could be type II GT where clot retraction can be normal or moderately diminished. Confirmation of the type would require analysis of GP IIb IIIa expression by flow cytometry. Hence, 4 of 15 patients (26.6%) in our study may be type II GT and would require flow cytometry for further confirmation.

Although clinical presentation and the screening tests help in clinching the diagnosis of GT, platelet aggregation studies are essential to confirm the diagnosis of GT and to rule out the other platelet function disorders. Thirteen patients in our study had platelet aggregation studies done and all of them showed decreased or absent aggregation with ADP, collagen, and epinephrine. This finding was consistent with the other studies. [2],[3] These agonists activate platelets through GP IIb-IIIa complex and hence in GT patients they fail to induce platelet aggregation.

Ristocetin interacts with platelets through GP Iba and Von Willebrand factor and does not require GP IIb-IIIa complex to cause platelet aggregation and hence GT patients usually show normal aggregation with ristocetin. Alan Nurden in his review article on GT has stated that normal ristocetin-induced platelet aggregation in GT helps to distinguish it from Bernard Soulier Syndrome. [1]

But various studies done on ristocetin-induced aggregation have shown that it may be affected in GT also. [14],[15],[16] The present study showed normal aggregation with ristocetin in seven patients; however, six patients showed decreased aggregation with ristocetin. Badhe

et al. [2] in their study also showed abnormal aggregation with ristocetin in three of their GT patients. Howard and Firkin in 1971 have also shown that platelet aggregation with ristocetin can be affected in GT patients. [14]

The exact reason why ristocetin-induced platelet aggregation is affected in GT is not known, but various hypothesis have been put forward to explain it. Studies on mechanism of ristocetin-induced platelet aggregation started way back in 1970s. Cohen et al. [17] in 1975 reported peculiar features of thrombasthenic platelets whereby ristocetin-induced platelet aggregation was spontaneously reversible and furthermore was inhibited by ADP. In their study, they noted that when ADP was added before ristocetin to thrombesthenic PRP, aggregation was inhibited, whereas addition of ATP following ristocetin or preceding ADP caused normal aggregation. ADP or ATP did not affect ristocetin-induced aggregation in normal subjects.

A likely interpretation for this observation could be that ADP binds to thrombesthenic platelets and causes a conformational change in ristocetin receptor and/or in the platelet membrane due to which the ristocetin receptor gets either buried or transformed only in case of GT platelets which are known to present various signs of abnormal membrane structure.

Chediak et al. [12] in 1978 have also suggested that the disaggregation phase of ristocetin response of thrombesthenic patients is not a function of the known GP membrane defect but depends on materials originating in the platelets. They noticed that control subject citrated PRP showed irreversible aggregation with ristocetin concentrations of 1-2 mg/ml whereas thrombestenic citrated PRP manifested several cycles of aggregation and disaggregation. By following the reaction for 6 minutes and by increasing the concentration of ristocetin of 1-2 mg/ml, they have observed a repetitive pattern of aggregation and disaggregation (four to six cycles) ending in partial disaggregation approximately 15-20 minutes after addition of R.

Mitsuru Murata has shown in their study done in 1991 that electronegative residues located in the carboxyl-terminal region of the 45-kDa domain of GP Ib are essential for the expression of vWF binding function. Decreased acidic residues impairs the binding GP Iba to Vwf even in the presence of Ristocetin. So in these cases, platelet aggregation studies will show reduced aggregation test with Ristocetin. [18] Defective ristocetin-induced platelet aggregation in GT could be related to all these factors.

Out of the six patients who had a decreased ristocetin-induced platelet aggregation in our study three also had a normal clot retraction. Hence, decreased platelet aggregation with ristocetin could be an association of type II GT where clot retraction can be normal. One of these three patients also had a normal bleeding time. This could be a variant form of GT. Hence, in these 6 (40%) patients who had unusual findings on laboratory investigations use of more advanced tools such as flow cytometry is required to confirm the diagnosis and to exclude other platelet functional defects.


   Conclusion Top


Glanzmann thrombasthenia has a high prevalence in southern India due to high incidence of consanguineous marriages in this region. Presenting at an early age GT patients usually present with mild to moderate mucocutaneous bleeding rarely requiring blood transfusions. However, females in reproductive age group can present with severe menorrhagia.

Increased BT with a normal platelet count, absent, or reduced clot retraction and absence of platelet clumps on a direct smear are the basic laboratory parameters which initiate a diagnosis of GT. The diagnosis is further supported by a normal coagulation profile. These findings could be variable in the type II and variant form of GT, hence platelet aggregation becomes essential in confirming the diagnosis of GT.

The classical pattern of platelet aggregation in GT is reduced or absent aggregation with all agonists (ADP, collagen, epinephrine) except ristocetin. However, ristocetin-induced aggregation can also be affected in GT in which cases differentiating it from other platelet disorders may become difficult. Therefore, use of monoclonal antibodies and flow cytometry to quantitate GP IIb-IIIa is becoming extensively popular in confirming diagnosis of GT and also identifying the type of the disease.

 
   References Top

1.Nurden AT. Review Glanzmann thrombasthenia. Orphanet J Rare Dis 2006;1:10.  Back to cited text no. 1
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2.Badhe BA, Jayanthi S, Dutta T. Clinical spectrum of Glanzmann thrombasthenia. Indian J Pathol Microbiol 2006;43:297-302.  Back to cited text no. 2
    
3.George JN, Caen JP, Nurden AT. Glanzmann thrombasthenia: the spectrum of clinical disease. Blood 1990;75:1383-95.  Back to cited text no. 3
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4.Afrasiabi A, Artoni A, Karimi M, Peyvandi F, Ashouri E, Mannucci PM. Glanzmann thrombasthenia and Bernard Suolier Syndrome in south Iran. Clin Lab Haematol 2005;27:324-7.  Back to cited text no. 4
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5.Nurden AT, George JN. Inherited abnormalities of platelet membrane. Hemostasis and Thrombosis. Basic principles and clinical practice. 5 th ed., chap. 66. Philadelphia: Lippincott Williams and Wilkins; 2005. p. 988.  Back to cited text no. 5
    
6.Ali N, Moiz B, Shaikh U, Adil S, Rizvi B, Rahman Y. Diagnostic tool for Glanzmann Thrombasthenia Clinicopathologic spectrum. J Coll Physicians Surg Pak 2008;18:91-4.  Back to cited text no. 6
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7.Khanduri U, Pulimood R, Sudarsanam A, Carman RH, Jadhav M, Periera S. Glanzmann thrombasthenia: A review and report of 42 cases from South India. Thromb Haemost 1981;46:717-21.  Back to cited text no. 7
    
8.Trasi SA, Pathare AV, Shetty SD, Ghosh K, Salvi V, Mohanty D. The spectrum of bleeding disorders in women with menorrhagia: A report from Western India. Ann Hematol 2005;84:339-42.  Back to cited text no. 8
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9.Ray AK. Nature, amount and extent of consanguinity among two South India castes. J Hered 1979;70:281-2.  Back to cited text no. 9
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10.Newman PJ, Seligsohn U, Lymen S, Coller BS. The molecular genetic basis of Glanzmann thrombastenia in the Iraqi, Jewish and Arab population in Israel. Proc Natl Acad Soc USA 1991;88:3160-4.  Back to cited text no. 10
    
11.Toogeh G, Sharifian R, Lak M, Safaee R, Artoni A, Peyvandi F. Presentation and pattern of symptons in 382 patients with Glanzmann thrombasthenia in Iran. Am J Hematol 2004;77:198-9.  Back to cited text no. 11
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12.Peerschke EI, Zucker MB, Grant RA, Egan JJ, Johnson MM. Correlation between fibrinogen binding to human platelets and platelet aggregability. Blood 1980;55:841-7.   Back to cited text no. 12
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13.Chediak J, Telfer MC, Vander Laan D, Maxey B, Cohen L. Cycles of agglutination and disagglutation induced by Ristocetin in thrombasthenic platelets. Br J Haematol 1979;43:113-26.  Back to cited text no. 13
    
14.Firkin BG, Howard MA, Farmer SJ. Observations on the ultrastructure of platelets in Glanzmann thrombasthenia. Br J Haematol 1974;27:527-31.  Back to cited text no. 14
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15.Weiss HJ, Roger J, Brand H. Defective Ristocetin induced platelet aggregation in Von Willebrand disease and its correlation by factor VIII. J Clin Investig 1973;52:2697-707.  Back to cited text no. 15
    
16.Howard MA, Firkin BG. Ristocetin--A new tool in the investigation of platelet aggregation. Thromb Diath Haemorrh 1971;26:362-9.  Back to cited text no. 16
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17.Cohen I, Glaser T, Seligsohn U. Effects of ADP nad ATP on bovine fibrinogen and ristocetin induced platelet aggregation in Glanzmann's thrombasthenia. Br J Haematol 1975;31:343-7.  Back to cited text no. 17
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18.Murata M, Ware J, Ruggeri ZM. Site directed mutagenesis of a soluble recombinant fragment of platelet glycpoprotein Iba demonstrating negatively charged residues involved in Von Willebrand factor binding. J Biol Chem 1991;266:15474-80.  Back to cited text no. 18
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    Figures

  [Figure 1], [Figure 2], [Figure 3]
 
 
    Tables

  [Table 1], [Table 2]


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