Compound heterozygosity for one novel and one recurrent mutation in a Thai patient with severe protein S deficiency.

Homozygous or compound heterozygous protein S (PS) deficiency is a very rare disorder in the anticoagulant system, that can lead to life-threatening thrombotic complications shortly after birth. This report describes the results of the genetic analysis of the PROS 1 genes in a Thai girl patient. She was reported in 1990 as the first case with homozygous PS deficiency and neonatal purpura fulminans. In the present report, we identified the mutations in this patient by direct sequencing of PCR products representing all 15 exons of the PROS 1 gene and their flanking intronic regions. The patient turned out to be compound heterozygous for two null mutations. One allele contained a novel sequence variation, an A-insertion in an A5-tract covering codon 146 and 147, that results in a frameshift and a stop codon (TAA) at position 155. The other allele contained a nonsense mutation in exon 12 by a transition at codon 410 CGA (Arg) to TGA (stop). Cosegregation of PS deficiency with these two genetic defects was observed in her family.

Bone marrow transplantation for thalassemia in Thailand.

We report our experience of bone marrow transplantation for thalassemia in Thailand. From July 1988 to September 1992, 10 thalassemic patients underwent allogeneic bone marrow transplantation. Two of them were homozygous beta-thalassemia and 8 were beta-thalassemia/Hb E disease. Seven were male and 3 female. The age ranged from 1.8-14 years. The conditioning regimen comprised busulfan 14 mg/kg and cyclophosphamide 200 mg/kg. For GVHD prophylaxis, either cyclosporine alone or in combination with short methotrexate was given. Five patients were alive and well 104-1534 days after transplantation. Three patients with severe manifestations at the time of transplant had partial engraftment, and lost their graft within 3 months. They survived with thalassemia 1041-1357 days after transplantation. One patient who received one antigen mismatched marrow from her brother had no engraftment and was alive with thalassemia 1429 days posttransplant. One patient died early on day 9 from CNS complication. No GVHD was observed in this series. These results indicate that bone marrow transplantation can cure thalassemia but there is still high autologous recovery rate in those with severe manifestations.

Successful bone marrow transplantation in a child with red blood cell pyruvate kinase deficiency.

We report the first successful use of BMT for the treatment of RBC pyruvate kinase (PK) deficiency in a boy who developed neonatal jaundice and severe transfusion-dependent hemolytic anemia a few months after birth. He received a BMT at the age of 5 from an HLA-identical sister who has normal PK activity after conditioning with busulfan and cyclophosphamide. The post-transplant course was uneventful. At present, 3 years after transplant, he is 8 years old and has a normal hemoglobin level and normal RBC PK activity without evidence of hemolysis. DNA analysis has confirmed full engraftment.

Bone marrow, peripheral blood and cord blood stem cell transplantation in children: ten years' experience at Siriraj Hospital.

Stem cell transplantations were performed in 69 children at Siriraj Hospital over a ten year period. The source of stem cells was bone marrow (60), peripheral blood (3), or cord blood (6). The diseases treated included 35 thalassemias, 11 Burkitt's lymphoma, five non-Hodgkin's lymphoma, five aplastic anemia, eight acute leukemia, and one each of neuroblastoma, severe combined immunodeficiency, Wiskott-Aldrich syndrome, myelodysplastic syndrome, and pyruvate kinase deficiency. The success rate of stem cell transplantation in Thai children varied according to the underlying diseases of the patients, ranging from 50% in acute leukemia to 100% in aplastic anemia. The outcome of stem cell transplantation in 35 thalassemic children revealed 23 (79.4%) were cured, whereas three (10.3%) remain alive with disease and the other three (10.3%) died. The incidence of graft-versus-host disease was low hen compared with that of Western countries. It is concluded that bone marrow, peripheral blood and cord blood stem cell transplantation will be the treatment of choice and will be widely used in the future to cure many hematologic and malignant disorders in children.

Vertical transmission of the hepatitis B surface antigen in Thailand.

The vertical transmission of the hepatitis B surface antigen (HBsAG) from HBsAg carrier mothers to their infants and children were studied in 42 mother-infant pairs, 27 siblings aged under 5 years and 34 fathers of these families. Thirteen out of 42 (30.9%) infants born from these mothers became HBsAg carriers at 3 to 6 months of age. Vertical transmission to their infants (76.5%) and other siblings (88.9%) occurred only in those mothers with e-antigen (HBeAg) positive but none in HBsAg carrier mothers without HBeAg. Most of the HBsAg carrier babies (70.5%) and their siblings (77.7%) born from the carrier mothers with HBeAg positive also had HBeAg in their sera. Good correlation of the presence of HBeAg and higher titer of HBsAg was found both in HBsAg carrier mothers and in their off-springs. This study clearly shows that HBsAg carrier Thai mothers with HBeAg transmitted hepatitis B virus vertically to their infants and children more readily than do the HBsAg carrier mothers without HBeAg.

Diagnostic value of buffy coat preparation in dengue hemorrhagic fever.

Examination of the buffy coat preparation is a simple, rapid diagnostic aid for the early diagnosis of dengue hemorrhagic fever (DHF). The presence of many transformed lymphocytes, usually 20-50 per cent during the first few days of fever is a unique finding especially during a secondary infection. Similar findings are absent during the early phase of illness in other viral infections. The buffy coat appearance in DHF is also clearly different from those in patients with bacterial infections, which show many polymorphonuclear leukocytes containing toxic granules and vacuoles. The early buffy coat diagnosis in 320 cases of DHF correlated well with the later serologic diagnosis. The buffy coat diagnosis of secondary DHF was correct in 94.2% of cases as early as the third day of fever. This technique has proven to be clinically useful for early screening test of secondary DHF in Southeast Asian countries. The observation of many transformed lymphocytes in the peripheral blood of DHF patients also offer some insight into the pathogenesis of this condition.

Kinetics of lymphocyte subpopulations in dengue hemorrhagic fever/dengue shock syndrome.

A kinetic study of lymphocyte subpopulations was performed in 61 dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS) patients aged 8 months to 12 years and in 59 age-matched normal controls. There were 36 patients in grade 2 and 25 patients in grade 3 of the disease severity. The studies were performed on febrile stage, the day of subsidence of fever or shock stage, 3 subsequent days after subsidence of fever or shock, and once on the recovery stage (approximately 14-18 days after subsidence of fever or shock). The study revealed that the absolute total lymphocytes, CD3+, CD4+, CD8+ and HNK-1+ cells were decreased on febrile stage and their lowest values were noted on the first day of subsidence of fever or shock, while B1+ cells were in the normal range. Thereafter, all lymphocyte subpopulations were increased. The total lymphocytes, B1+ and CD8+ cells were rapidly increased and were above normal value on day 2 after subsidence of fever or shock (early convalescence), then gradually declined to the normal range. In contrast, CD3+, CD4+ and HNK-1+ cells were increased gradually and reached their normal values on day 2 after subsidence of fever or shock. The T4:T8 ratio began to reverse on the day of subsidence of fever or shock, reached its peak on day 2 after shock and returned to normal ratio rapidly thereafter. Thus, the absolute lymphopenia on the day of shock was due to the decrement or T cells (both CD4+ and CD8+ cells) and HNK-1+ cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Hemophilia with factor VIII and factor IX inhibitors, incidence, bleeding problems and management.

Among 117 cases of hemophilia, there were 7 hemophilia A and 2 hemophilia B with factor VIII and factor IX inhibitors diagnosed at the Department of Pediatrics, Siriraj Hospital, Bangkok, Thailand. The overall incidence of hemophilia with inhibitors was 7.7%. Eight cases (6 hemophilia A. 2 hemophilia B) were severe hemophilia and 1 moderate hemophilia A. The average age of the inhibitor detection was about 5 years. Of the 9 cases, 7 had high inhibitor titers and 2 had low inhibitor titers. The frequency of bleeding problems before and after inhibitor detection were not different. The bleedings included hemarthrosis, mucosal bleed, hematoma, oozing from wound, hematuria and intracranial hemorrhage. The treatment of hemarthrosis in hemophilia A with low inhibitor titers was the combination of short course of prednisolone and single large dose factor VIII. In high inhibitor titer patients with acute hemarthrosis (both hemophilia A and hemophilia B), the treatment consisted of prednisolone short course and single high dose of PCC. For bleeding control in both high and low inhibitor titer with mucosal bleeds, oozing from wounds, central nervous system bleeding and hematuria, the combination was used of high dose factor VIII or factor IX for 2 days, and tranexamic acid, prednisolone, cyclophosphamide were required. In life-threatening hemorrhage and surgical operation, plasmapheresis and large dosage factor VIII or factor IX were the treatment of choice. All supportive measures were also important in every case of mucosal bleeds, wounds and surgical operations. The result of treatment revealed one death from massive intracranial hemorrhage and 8 survivals, with joint contracture in 2 cases. All still have inhibitor detected, but in low titer.

Detection of varicella-zoster antibody by immune adherence hemagglutination test.

The immune adherence hemagglutination (IAHA) is a more sensitive and specific test for evaluation of humoral immunity to varicella-zoster virus (VZV) than a conventional complement fixation (CF) test. The equipment, reagents, procedure used in the IAHA are basically the same as those employed in the CF test. After VZ vaccination, seroconversions detected by IAHA test were significantly greater than those by CF test (P less than 0.01, chi 2 test). Moreover, the IAHA titers were higher about 4 times (G.M. 30.9 vs 8.2) and persisted longer than CF titers. Although the IAHA antigen is not a commercial product, the IAHA is sensitive and suitable assay for VZ antibody determination in routine serological laboratory.

Evidence of vitamin K deficiency in cord blood.

The prevalence of vitamin K deficiency in the newborns delivered at Siriraj Hospital was studied. The prolongation of one stage prothrombin time and the presence of PIVKA-II (non carboxylated prothrombin antigen) in cord blood were interpreted as the secondary change from vitamin K deficiency state. The most reliable method to diagnose vitamin K deficiency is the detection of vitamin K level in plasma which is not yet available in Thailand. Although the prevalence of vitamin K deficiency in the newborns from our data is not high, only 0.6%, it is shown that some of the apparently normal newborn infants may have bleeding problem from vitamin K deficiency in both newborn and early infancy periods. So, the correction of this deficiency by administration of vitamin K to all newborns is appropriate and reasonable decision.

Abnormal hemostasis in dengue hemorrhagic fever.

Abstract. Abnormal hemostasis in dengue hemorrhagic fever includes:- 1. Vasculopathy which occurs during the early febrile to pre-shock and shock phase. The evidences support are: 1.1 Increased anaphylatoxin, released by complement activation causing leakage of intravascular fluid in to serous space. 1.2 Positive tourniquet test, some of which occur preceeding thrombocytopenia in the acute phase of DHF. 1.3 Excessive increased in PGI2 which is the most potent vasodilator and platelet aggregation inhibitor. 2. Platelets: 2.1 Thrombocytopenia due to 2.1.1 The bone marrow hypocellularity with increased in all forms of megakaryocytes but the vacuolated and disintegrated ones. 2.1.2 Destruction by the liver and spleen. 2.1.3 Immune-mediated injury as demonstration of dengue antibody complexes on the platelet surface. 2.1.4 The in vitro spontaneous aggregation to vascular endothelial cell pre-infected by dengue virus inducing platelet aggregation, causing lysis and platelet destruction. 2.2 Dysfunction shown by 2.2.1 Increased release of betathromboglobulin (BTG), PF4 and PGI2. 2.2.2 In vitro hypoaggregation stimulated by ADP and defect in ADP-releasing ability. 3. Coagulopathy including: 3.1 Prothrombin complex deficiency due to liver damage. 3.2 Consumptive coagulopathy due to the activation by mononuclear phagocytes, PF3 released from platelet aggregation. DIC is seen in prolonged shock cases of DSS.

Transient, acquired glucose-6-phosphate dehydrogenase deficiency in Thai children with typhoid fever.

Seventy nine children with typhoid fever were studied, 45 were males and 34 were females. There were 9 out of 45 males patients (20%) who had definite G-6-PD deficiency of whom 3 had acute intravascular hemolysis. The rest of the patients had transient low enzyme activity during the first few weeks of their illness, with reticulocytopenia. Their G-6-PD activities rose up to normal level later in the course of the disease while the reticulocytes were also increased. This study demonstrated that even in normal G-6-PD subjects, typhoid fever can cause transient, acquired low G-6-PD level due to bone marrow suppression. It was suggested from this study that quantitative G-6-PD assay was more useful and sensitive than the screening method and that long term follow up is needed in the case that had unexplained low G-6-PD activity.

Susceptibility to varicella-zoster virus in Thai children and young adults.

During 1982-1983, susceptibility to varicella-zoster virus (VZV) in 224 Thai subjects at high risk for varicella infection was studied. The immune adherence hemagglutination (IAHA) and VZ skin test were carried out to determine VZV immunity in immunocompromised children and young adults. The history of varicella and herpes zoster from each subject was recorded. The mean +/- SD age in children and young adults were 7.3 +/- 2.8 and 19.6 +/- 1.2. Negative IAHA test was found in 74.2% of 62 children and 35.2% of 162 young adults. The increase in immune individuals was demonstrated with advancing age. Response to VZ skin test showed positive results in 79 of 162 (48.8%) young adults. The seronegativity was related to the negative VZ skin test (p less than 0.001, X2 test). Regardless of antibody detection or VZ skin test, 47 of 162 (29%) young adults were susceptible. According to the positive history of varicella and of herpes zoster obtained from 95 young adults, 80% had developed varicella during 1 to 10 years of age and 8.8% had positive history of herpes zoster. The findings suggest that the IAHA and VZ skin test should be used together for assessing VZ immunity. Varicella vaccination is highly recommended for susceptible persons who may develop severe illness.

Study of hematopoietic progenitor cells in thalassemia after bone marrow transplantation.

The hematopoietic committed progenitor cells (BFU-E and CFU-GM) in blood and bone marrow were studied in thalassemic patients before and after bone marrow transplantation. Eighteen transplants were performed in 17 patients with thalassemia. Five were homozygous beta-thalassemia and 12 were beta-thalassemia/hemoglobin E disease. The age ranged from 1.2-14 years (median = 3.4 years). The conditioning regimen comprised busulfan 3.5-4 mg/kg/day for 4 days and cyclophosphamide 50 mg/kg/day for 4 days. Cyclosporin in combination with methotrexate were administered post transplant for GVHD prophylaxis. Before transplantation, BFU-E and CFU-GM in the blood of the patients were significantly higher compared with normal (p < 0.05) but were normal in the bone marrow. Only the CFU-GM in the bone marrow of the successful cases after transplantation recovered to the normal level at the first month through the 12th month whereas the BFU-E were low. Both CFU-GM and BFU-E in the blood were lower than normal after follow up to the 12th month. Inspite of the low number of progenitor cells, there was hematological recovery in the blood of the patients. It may be due to the capacity of the hematopoiesis react to stress which could be amplified the differentiation compartment or the shortened-transit time through the stem cell compartment.

Risk of transfusion associated AIDS by seronegative blood in Thailand: a multicenter report.

Prevention of transfusion associated AIDS (TAA) in Thailand began in 1986 when the HIV infection started to be sharply increased among the general population. The retrospective anti-HIV screening in various blood donor populations by The National Blood Center (NBC) revealed a seroconverted prisoner. Then the use of prisoners, prisoners' blood was not recommended from 1986. In April 1987, the first case of TAA was disclosed. Five months later, anti-HIV screening in all units of blood was firstly introduced at Ramathibodi Hospital (RH) and NBC. From 1989, anti-HIV screening in all units of blood is mandatory nationwide by Ministry of Public Health. Despite the anti-HIV screening, TAA cases transmitted by seronegative blood were gradually reported. Among many Medical Centers, there were 9 and 18 cases of TAA recorded from Chiang Mai and Bangkok areas respectively, since 1985. In addition, several new seroconverters were observed among voluntary blood donors. All of this evidence indicates the existence of blood donation during the early stage of infection, the so-called "window period". At present, HIV-P24 antigen ELISA seems to be the only available technique for mass screening. In 1990, NBC successfully performed a retrospective study on HIV-Ag ELISA screening by obtaining the prevalence of 1/10,000 units of blood. At the same period of time, in RH prospective study, a unit of blood with HIV-Ag only was detected when 3432 units of blood were screened. The HIV-Ag ELISA screening was then performed on every unit of blood routinely since Aug 12, 1991 at RH.(ABSTRACT TRUNCATED AT 250 WORDS)

Burkitt's lymphoma in Thai children: an analysis of 25 cases.

Twenty five cases of Burkitt's lymphoma in Thai children were diagnosed at the Department of Pediatrics, Siriraj Hospital during the period of 13 years (January 1969 to April 1982). Males were more affected than females with the ratio of 1.7:1. The age ranged from 2 to 11 years with the median age of 4-5 years. The most common clinical manifestations were abdominal mass associated with nausea vomiting, abdominal pain, anorexia, weight loss and generalized lymphadenopathy which occurred in 50-60% of cases. Additional symptoms and signs included anemia, hepatosplenomegaly, edema and pleural effusion. Jaw tumor was found in only 37.5% of the patients. Definite diagnosis depended on the characteristic starry sky appearance of the lymph node biopsy or section of abdominal mass. In advance cases, the tumor cells could be discovered in bone marrow aspiration, ascitic fluid pleural fluid and cerebrospinal fluid. The typical blast cells were detected in the peripheral blood in 4 cases. Antibody to Epstein-Barr virus could be detected in almost all cases with high titers in some cases. Most patients responded very well to local irradiation and chemotherapy with prednisolone plus cyclophosphamide and vincristine or methotrexate. However, relapse occurred rapidly and 80% of the patients died within 3 months after diagnosis with the median survival of only 1 month. Five cases expired early before any specific treatment. The main causes of death were disease, sepsis, excessive bleeding and hyperkalemia.(ABSTRACT TRUNCATED AT 250 WORDS)

Niemann-Pick disease in association with homozygous hemoglobin E: a case report.

The first case of Niemann-Pick disease Type A in a Thai infant was reported. The patient also had abnormal hemoglobin E. The diagnosis was based on the clinical features, bone marrow findings and sphingomyelinase levels in the culture of skin fibroblasts. The autosomal recessive mode of inheritance was confirmed in this case.

Pyruvate kinase deficiency in an alpha-thalassemia family: first case report in Thailand.

In Thailand, the most common cause of chronic hemolytic anemia is thalassemia hemoglobinopathy. We report here a 10-year-old girl with pyruvate kinase (PK) deficiency who was initially diagnosed to have Hb H disease, like her sister. The patient had a history of neonatal jaundice which required blood exchange transfusion twice and phototherapy. She became anemic and regular blood transfusion was required since the age of 2 1/2 months. She was very anemic compared to her sister and was transfusion dependent. Besides, she never had red cell inclusion bodies, thus re-evaluation was performed. The diagnosis of red cell pyruvate kinase deficiency and the exclusion of Hb H disease was achieved after cessation of blood transfusion for 3 months. The family study also confirmed the diagnosis. The patient is now on high transfusion and iron chelation. She is doing well with mild splenomegaly.

Successful prophylaxis of intracranial hemorrhage in infants with severe congenital factor VII deficiency.

During the period 1984-1992, 2 severe cases (1 male, 1 female) of congenital F VII deficiency with intracranial hemorrhage (ICH) were referred to the Department of Pediatrics, Siriraj Hospital Bangkok, Thailand at the ages of 1 and 3 months old. They both responded very well to fresh frozen plasma (FFP) transfusion therapy. Subsequently, both had repeated episodes of ICH (repeated ICH) 5 and 6 times, despite the 10-14 days of replacement therapy for each episode and eventually died at the ages of 11 and 13 months. Since September 1996, another 2 severe cases (2 females) of congenital F VII deficiency who had ICH within their first month of life were referred to us. In order to prevent repeated ICH, we started a prophylactic regime after the second episode of ICH, by giving FFP 10 ml/kg twice a week. The average duration of follow up was 21 months (at 8 and 34 months). All of them (aged 14, and 38 months old) are doing well at this time and free from repeated ICH. From this observation, if there is FFP available, this regime is an effective way to prevent repeated ICH in infants with severe congenital Factor VII deficiency.

Glucose-6-phosphate dehydrogenase deficiency in the newborn: its prevalence and relation to neonatal jaundice.

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is prevalent in Thailand. This condition can cause acute hemolysis during oxidative stress and also severe hyperbilirubinemia in the newborn in some populations. Our aim was to study the prevalence of G6PD deficiency in relation to neonatal jaundice. We performed quantitative red blood cell (RBC) G6PD assay in the cord blood of 505 male subjects. Observation of jaundice and determination of bilirubin level as well as work up for other causes of jaundice were made in the G6PD deficiency group compared to a G6PD normal group. Questionnaires were also sent for further follow up to both groups. The results of the study were as follows: Sixty-one of 505 male (12.08%) had RBC G6PD deficiency (Group I). The rest (444 cases) had normal G6PD (Group II). In Group I, 49.15% developed neonatal jaundice, of which 28.82% were physiologic and 20.33% were pathologic jaundice. In group II, 23.68% developed jaundice; 16.51% were physiologic and 7.17% were pathologic jaundice, respectively. Onset of jaundice, date of peak bilirubin and peak bilirubin level in Group I and Group II were not statistically different. ABO incompatibility was associated with Group I in 17.24% and with Group II in 9.09%. Hospitalization day in Groups I and II were not statistically different. Other associated diseases were found in both groups, ie infection, congenital malformation, respiratory distress syndrome, but there was no significant difference in terms of jaundice. Phototherapy was required in 18.64% and 10.28% in Group I and II with a duration of 3.91 +/- 1.24 and 3.21 +/- 1.75 days, respectively. One case in Group I who was also premature received one exchange blood transfusion due to severe sepsis but he did not survive. One case in Group II who had polycythemia was successfully treated by partial exchange transfusion with plasma.