Laboratory Diagnosis of Congenital Factor V Deficiency, Routine, Specific Coagulation Tests with Molecular Methods

Erratum: The correct affiliation of corresponding author of this manuscript has been edited as follows:"Akbar Dorgalaleh: Department of Hematology and Blood Transfusion, School of Allied Medical Sciences, Iran University of Medical Sciences, Tehran, Iran."Factor V (FV) deficiency is a rare bleeding disorder (RBD) that inherit in autosomal recessive manner. Diagnosis of FV deficiency (FVD) is made by routine coagulation tests, FV activity and molecular analysis. In patients with FVD, routine coagulation tests including activated partial thromboplastin time (APTT), prothrombin time (PT) and evenbleeding time (BT) are prolongedwhile thrombin time (TT) is normal. FV activity assay can use for confirmation of diagnosis as well as for differential diagnosis with acquired forms of disease. Mixing study can be used for screening of inhibitor against FV. In this situation, addition of normal plasma cannot correct prolonged PT and PTT while in congenital FVD prolongation is corrected. Molecular diagnosis of FVD is straightforward but due to large size of FV gene and genetic variability molecular diagnosis is restricted to research laboratory

Burden of Congenital Factor XIII Deficiency in Iran

Congenital factor XIII (FXIII) deficiency is a rare coagulopathy with the highest incidence in Iran. Iranian patients with FXIII deficiency (FXIIID) presented high rate of bleeding episodes, some of them are major cause of disability and mortality among these patients. Hemarthrosis and intracranial hemorrhage (ICH) can affect activity and social productivity of patients. ICH, recurrent miscarriage and umbilical cord bleeding are the major cause of mortality. Hematoma, and prolonged menstrual bleeding as well as post-surgical bleeding are other significant bleeding in Iranian patients with FXIIID. Present of severe life threatening bleeding episodes and other notable bleedings, can significantly reduce working activities and social productivities of patients. Although Iranian patients with FXIIID, experienced significant diseases related complications, early diagnosis accompany by appropriate therapeutic regimes can prevent most of these problems

c.559 T>C as The Most Common Mutation of Factor XIII Deficiency in Iranian Patients is not Restricted to Southeast Iran

Background: Iran has a large group of patients with severe congenital factor XIII deficiency (FXIIID) and Trp187Arg mutation that is most disease causing mutation of FXIII in the world is only observed in southeast of Iran with 352 patients with FXIIID. 743 patients with FXIIID was observed in 17 provinces of Iran but Tehran city with more than 12 million population has no any registered patient with FXIIID. Here we described first case with severe congenital FXIIID in Tehran Province with underline FXIII mutation.Methods: A neonate with prolonged umbilical cord bleeding was referred to hemophilia center. Patient was screened by routine coagulation tests and by clot solubility test. After observation of normal routine tests and abnormal clot solubility patient was underwent a full sequencing of FXIII-A gene. For confirmation of detected mutation in FXIII-A gene, exon 4 was amplified by PCR and cleaved by Eco130I restriction enzyme.Results: We found first case with severe congenital FXIIID in Tehran Province with Trp187Arg mutation in exon 4 of FXIII-A gene. Patient’s parents were heterozygote for this mutation.Conclusion: Trp187Arg mutation of FXIII-A is the most common mutation in Iranian patients with FXIIID and is not restricted to southeast of Iran.Keywords: Factor XIII deficiency, Trp187Arg mutation, Tehran Provinc

Coagulation Factor XIII-A A614T gene Variation is Suggestive of Founder Effect in Iranian Patients with Severe Congenital Factor XIII Deficiency

Background: Factor XIII (FXIII) is a heterotetramer consisting of two subunits, FXIII-A and FXIII-B. Several common gene variations were observed in the FXIII-A gene with an obvious ethnic difference. This study assessed the prevalence of A614T as a common FXIII-A gene variation among Iranian patients with FXIII deficiency (FXIIID). Materials and Methods: This study was conducted on eighty Iranian unrelated individuals with FXIIID. Genotype analysis for FXIII-A A614T gene variation was performed for all individuals. Results: Molecular analysis of these Iranian populations revealed that all studied patients were homozygous for the T allele at codon 204 of the FXIII-A1 subunit. Conclusion: Present of T allele at codon 204 of FXIII-A1 subunit among all study population can be suggestive of founder effect. &nbsp

Genetic Risk Factors for Inhibitor Development in Patients with Hemophilia and Rare Bleeding Disorders

Inhibitor development is a lifelong challenge for patients with bleeding disorders who received replacement therapy. Most commonly, inhibitor formation was observed in hemophilia A patients but patients with rare bleeding disorders (RBD) especially patients with deficiency of factor XIII (FXIII) and factor V (FV) can develop an inhibitor against exogenous factors. Several factors considered as risk factors for inhibitor formations in these patients. Genetic risk factors are the main accused that can cause inhibitor formation in hemophilia patients but are less important in RBDs.For this review study, we searched MEDLINE and Web of Science databases for English sources and the following key words: hemophilia, inhibitor, rare bleeding disorder, rare inherited disorder, acquired hemophilia, acquired rare bleeding disorders, treatment complication, genetic in hemophilia, polymorphism in rare bleeding disorder, mutation in hemophilia and other required keywords.Hemophilia A (HA) patients who had large deletion, nonsense mutation or intron 22 inversion are more susceptible for inhibitor development. Gene polymorphisms in immune system are also considered as other risk factors in HA patients.Keywords:  Hemophilia, rare bleeding disorder, inhibito

Congenital Prothrombin Deficiency

Congenital prothrombin deficiency is an extremely rare hemorrhagic disorder with estimated prevalence of 1 per 2,000,000 in the general population. Since the disorder is an autosomal recessive disorder, the disorder is more frequent in areas with high rate of consanguinity. Clinical manifestations of disorder are highly variable ranging from mild bleeding episodes to severe life-threatening hemorrhages. The disorder can be diagnosed based on routine and specific tests. No specific factor II concentrate is available, but patients can receive fresh frozen plasma and prothrombin complex concentrate (PCC). Traditionally patients with prothrombin deficiency receive on-demand therapy, but secondary prophylaxis can be used for those patients with high risk of severe life-threatening bleeding. With timely diagnosis and appropriate management of disorder, the quality of life in these patients can significantly improve.   Keywords: Prothrombin deficiency, Clinical manifestations, Diagnosis, Treatmen

Extensive Hematoma in a Patient with HereditaryHypersegmentation of Neutrophils

 Erratum: The correct affiliation of corresponding author of this manuscript has been edited as follows:"Akbar Dorgalaleh: Department of Hematology and Blood Transfusion, School of Allied Medical Sciences, Iran University of Medical Sciences, Tehran, Iran."Hypercoagulable states are a group of conditions associated with an enhanced tendency toward blood clotting. Although usual clinical manifestations of hypercoagulable states are thrombotic events such as deep venous thrombosis, hematoma can also occurs as a result of hypercoagulability in some patients. Several inherited or acquired conditions may lead to hypercoagulable states. Some of them include myeloproliferative syndromes, over activity of coagulation factors and methyltetrahydrofolateee reductase (MTHFR) polymorphisms. MTHFR is required for convertingthe amino acid   homocysteine to methionine. Another significant role of an aptly functioning MTHFR enzyme is nucleic acid biosynthesis. Therefore MTHFR polymorphisms are expected to be associated with hypersegmentation of neutrophils because of a defect in DNA metabolism. Neutrophil hypersegmentation is one of the most sensitive haematological features of cobalamin or folate deficiency with normal serum vitamin B12-folic acid and iron levels. Hypersegmentation of neutrophils and hematoma which  both of them suspected to be  due to gene variations of MTHFR. Here we report a 37 years old female who simultaneously affected by hereditary hypersegmentation and extensive hematoma. Laboratory analysis revealed normal serum vitamin B12, folic acid and iron levels. Routine and specific coagulation tests were normal in except of factor VIIIc that was high. Results of complete blood cell count (CBC) test were normal. Although this is just an idea, but simultaneous presentations of these two conditions can have a common origin

Gastrointestinal bleeding in a newborn infant with congenital factor X deficiency and COVID-19—A common clinical feature between a rare disorder and a new, common infection

Dear Editors, Congenital factor X (FX) deficiency is an extremely rare, bleeding disorder with an estimated incidence of one per 1 million. Patients with severe FX deficiency (FX:C < 1%) demonstrate a wide spectrum of serious clinical presentations, including hemarthrosis, hematoma, gastrointestinal (GI) bleeding, intracranial hemorrhage (ICH), and umbilical cord bleeding.1 In fact, severe FX deficiency, with a high rate of life‐threatening bleeding, is the second‐most severe, rare coagulation factor deficiency (RCFD) after FXIII deficiency.1, 2 Although homozygotes are at risk of severe bleeding, heterozygotes usually are asymptomatic, but postsurgical bleeding or bleeding after childbirth may occur.1, 2 Other risk factors can increase the risk of bleeding in FX deficiency, and coronavirus disease 2019 (COVID‐19), a new medical challenge, could affect the patient's bleeding or thrombotic tendency.3 COVID‐19, which is caused by severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) presents an enormous challenge for everyone, especially for those with underlying risk factors such as cardiovascular disease, diabetes, obesity, and renal failure. Age and male sex are other risk factors.4 Limited data are available regarding the effect of COVID‐19 on patients with congenital bleeding disorders (CBDs), particularly RCFDs.5 It has been shown that hypercoagulability‐related adverse consequences are less common among patients with CBDs, at least in those with moderate‐to‐severe deficiency, but further studies, including our ongoing work on a large number of patients, are required.5 Although there are several reports of newborns among infected pregnant mothers, this is the first report of such a case in an RCFD. This case report may help medical professionals to better manage similar cases. A 19‐year‐old pregnant woman was infected with SARS‐CoV‐2 early in the 9th month of pregnancy. Reverse transcriptase‐polymerase chain reaction (RT‐PCR) confirmed the infection. The patient had been in close contact with family members with confirmed COVID‐19. The patient had cough and fever. Due to the mild presentation, she was given Azithromycin and advised to isolate herself at home. The symptoms resolved within 14 days. At end of her 9th month, three days prior to the planned cesarean section, she was rechecked for SARS‐CoV‐2 infection; her RT‐PCR was negative. She successfully underwent cesarean section without complications and delivered a healthy full‐term baby. Therefore, mother and newborn discharged the following morning. In the evening, the baby experienced bloody vomiting and was hospitalized for further assessment, which showed GI bleeding. At admission, laboratory tests showed a positive C‐reactive protein (CRP) (qualitative), a low hemoglobin level, and prolonged prothrombin time (PT), and activated partial thromboplastin time (APTT) (Table 1). He was hospitalized in the neonate intensive care unit (NICU) for 10 days. Due to the risk of SARS‐CoV‐2 infection, on the third day after admission he was tested by RT‐PCR, which was positive. The neonate received 30 mL frozen plasma (FFP) six times over 10 days, which resolved the GI bleeding. Tranexamic acid (TXA) was administered at a dose of 10 mg/kg every 8 hours. Due to lack of COVID‐19 symptoms, he did not receive any special treatment for the disorder. After 10‐day hospitalization in the NICU, the neonate was sent to an isolation room for 5 days, during which his condition stabilized, after which he was discharged in stable condition. He has had no complications during the past two months after discharge. Since the child's father and two other first‐degree family members have severe FX deficiency, and the parents of the baby are closely related, the mother and the baby were checked for FX deficiency. Routine coagulation tests, and FX:C assay performed by STA Compact automatic coagulometer (Stago, Paris, France), revealed a severe deficiency in the baby, and a mild deficiency, compatible with heterozygote FX deficiency, in the mother (Table 1). Table 1. Laboratory characteristics of mother and baby with factor X deficiency and COVID‐19 Test Proband (2nd day after birth) Proband (7th day after birth) Proband (2 months after hospital discharge) Mother (about 3 1/2 months after SARS‐CoV‐2 infection) WBC × 109/L 14.2 (8‐24)b 9.43 (5‐21) 10.79 (6‐18) 8.7 (3.6‐10.6) RBC × 109/L 2.5 (4.36‐5.96) 2.78 (4.2‐5.8) 3.50 (3.4‐5) 4.41 (3.8‐5.2) Hb (g/dL) 8.2 (16.4‐20.8) 9.2 (15.2‐20.4) 10.2 (10.6‐16.4) 13.6 (12‐15) HCT (%) 24.6 (48‐68) 27 (50‐64) 29.2 (32‐50) 41.4 (35‐49) Lymphocyte × 109/L 6.4 (1.3‐11) 4.3 (1.2‐11.3) 8.21 (2.5‐13) 2.22 (1‐3.2) Neutrophil × 109/L 4.9 (2.6‐17) 2.9 (1.5‐12.6) 1.85 (1.2‐8.1) 5.75 (1.7‐7.5) Platelet × 109/L 370 (150‐450) 331 (150‐450) 334 (150‐450) 276 (150‐450) PT (sec) >60 (PTC: 12.6) 90 (PTC: 12.6) >60 (PTC: 10) 13 (PTC: 10) APTT (sec) >120 (APTTC: 31) 100 (APTTC: 30) >120 (APTTC: 32) 37 (APTTC: 32) CRP (Quantitative) Trace Negative NC NC FX:C level NC NC <1% (50%‐150%) 40% (50%‐150%) Abbreviations: APTT, activated partial thromboplastin time; APTTC, APTT control; CRP, C‐reactive protein; Hb, hemoglobin; HCT, hematocrit; NC, Not checked; PT, prothrombin time; PTC, PT control; RBC, red blood cell; WBC, white blood cell. a Hematological test normal ranges are extracted from Rodak's Hematology: Clinical Principles and Applications, 5th Ed (2016). b Normal values are placed in parentheses. COVID‐19 is an emerging medical challenge that can present more difficulties for those with special conditions, such as pregnant women and newborns. Due to alterations in cellular immunity, pregnant women are more prone to infection by intracellular pathogens like viruses.6 The fetus is also highly susceptible to infection due to immaturity of the immune system.7 Furthermore, the mother's (heterozygote) congenital coagulopathy and that of her newborn (homozygote) were additional potential risk factors, because a disrupted coagulation system is a prominent feature of SARS‐CoV‐2 infection.8 To date, FX deficiency in a newborn has not been cited anywhere as a special condition requiring close attention in the case of SARS‐CoV‐2 infection. According to the few reports to date, SARS‐CoV‐2 infection is a risk factor for severe maternal morbidity. It is worth noting that most of those mothers were discharged without complications.9 From a clinical aspect, fever was the most common symptom (68%) at the time of admission.9 This was also observed in the affected woman of this study. SARS‐CoV‐2 infection can even affect the type of delivery. A systematic review of these women showed that about 92% of deliveries were by cesarean section, less than 10% being the usual vaginal delivery (7 of 85). Fetal distress was mentioned as the most common indication for cesarean section. Our patient underwent a planned cesarean section, due to her previous history. The delivery itself was uneventful, and a healthy baby was delivered, while among other reported cases, a number of complications have been noted.9 As with most other reports, the infant did not have any symptoms at the time of delivery and was discharged the day after birth.9 In a case series of 10 patients, various first clinical presentations were observed, including shortness of breath (n = 6), fever (n = 2), vomiting (n = 1), and rapid heart rate (n = 1).10 In the case at hand, bloody vomiting was the first clinical presentation. In the same case series, one died due to refractory shock, multiple organ failure (MOF), and disseminated intravascular coagulation (DIC). Another patient with severe presentation was managed by intravenous infusions of gamma globulin, platelets, and plasma, which was suggestive of the effectiveness of gamma globulin in severe cases. The author recommended early use of intravenous gamma globulin for passive immunization.10 GI bleeding in our case was successfully managed by administration of FFP and TXA. In addition to thrombotic complication, bleeding is not infrequent in patients affected by COVID‐19, with GI bleeding seemingly the most common hemorrhagic manifestation among adults. GI bleeding, with a frequency of 40%, was observed among neonates from affected mothers.3 On the other hand, GI bleeding is also a relatively common presentation among severely FX deficient patients.1, 2 In fact, GI bleeding can occur in children with severe FX deficiency within the first months of life. It seems that such patients are prone to experience severe bleeding, such as ICH, later in life, in the absence of an appropriate therapeutic strategy, most likely preventative regular secondary prophylaxis.1, 2 In one study of 102 patients with congenital FX deficiency, GI bleeding has been reported in 12% of symptomatic cases.1 In this case, with GI bleeding being a common presentation of SARS‐CoV‐2 infection and congenital FX deficiency, it cannot definitively be attributed to one or the other. Close monitoring of such cases is necessary to decrease related adverse consequences. Although it seems that COVID‐19 is less severe in adults with CBDs, it is a less‐known issue among children and newborns with CBDs. Further reports and studies could provide clarity. Due to their severe bleeding tendency, close monitoring of patients with severe congenital FX deficiency is mandatory, even without potential SARS‐CoV‐2 infection. And close monitoring of neonates with infected mothers is mandatory to prevent severe consequences. Patients with concomitant infection with SARS‐CoV‐2 require even more rigorous preventative and supportive care. ACKNOWLEDGEMENTS We highly appreciate Daisy Morant's valuable aid in improving the English Language of this manuscript. The study was supported and approved by Shahid Beheshti University of Medical Sciences. CONFLICT OF INTEREST The authors have no competing interests. AUTHOR CONTRIBUTIONS A. Dorgalaleh designed the work, performed laboratory analysis, and wrote the manuscript. F Ghazizadeh, M. Baghaipour, A. Dabbagh, Gh. Bahoush, and N Baghaipour performed clinical studies. Sh. Tabibian, M. Jazebi, N. Baghaipour, M. Bahraini, A. Fazeli, and F. Yousefi performed laboratory analysis. All the authors approved the submission

The effects of Hyperglycemia and Hyperlipidemia on blood indices

Background: Cell Blood Count (CBC) are performed with automatic analysers in laboratories.It works based on three principles electrical impedance, scatter light and flowcytometry. Many factors might affect results by these machines. This study was performed to assay the effect of some confounders on the results of analysers that work based on electrical resistance. Material and method: This analytical study (case- control) was conducted on 243 persons with hyperglycemia and hyperlipidemia and 100 healthy persosn (control group) in Fateme Zahra Hospital in North of Iran, Golestan. First, Blood glucose and triglyceride were measured with biochemical analyser and CBC was performed for each person. Finally, Statistical analysis and comparison between two groups were performed with SPSS software. Resuts: Comparison between case and control group was shown that both hyperlipidemia and hyperglycemia cause increase in&nbsp; Mean Cell Volume (MCV) and also hyperlipidemia can cause increase in Mean Cell Hemoglobin Concentration (MCHC), Hemoglobin and Hematocrit (Hct) with (P<0.05) Conclusion: According to this study, confounding factors such as hyperglycemia and hyperlipidemia can affect the results of analysers that work based on electrical impedance and it should be considered in laboratories