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Sentot Santoso

Publications and source records attributed to Sentot Santoso.

At least 19 recordsLinked to original sources
PubMed2007

A prospective study on the prevalence and risk factors for neonatal thrombocytopenia and platelet alloimmunization among 9332 unselected Brazilian newborns.

BACKGROUND: Neonatal thrombocytopenia (NT) occurs in 0.5 to 0.9% of unselected Caucasian newborns. However, the prevalence of this complication in other populations is unknown. In this study the prevalence/causes of NT was determined in Brazilian newborns, a population characterized by admixture among Indigenous, Africans, and Caucasians. STUDY DESIGN: A prospective study was carried out in a 3-year period, to determine the prevalence and causes of thrombocytopenia in cord blood samples. Genotyping for HPA 1-5 systems was performed in pairs of mother/neonates with and without thrombocytopenia. All mothers with genotypic mismatches from each group were tested for HPA-specific antibody using the MAIPA technique to identify alloimmunization. RESULTS: Platelet counts <100 x 10(9)/L were detected in 1.5% of 9,332 unselected newborns. In 0.15%, platelet count was <50 x 10(9)/L. Clinically significant bleeding was rare. Underlying diseases were present in 48% of the thrombocytopenic cases. HPA 1-5 system genotype mismatches occurred in 50% of gestations, but did not predict the risk for thrombocytopenia. Notably, mismatched genotypes for HPA-5 were slightly increased in the thrombocytopenic group. The presence of anti-HPA-5b antibodies was observed in 0.05% of unselected pregnancies, but increased to 12% among mothers of neonates with thrombocytopenia and mismatched genotype (N = 51). CONCLUSIONS: Neonatal thrombocytopenia is common among Brazilian newborns at rates comparable with those described among Caucasians. These data suggest that screening for genotypic HPA mismatch, followed by an HPA-specific immunoassay system, particularly for the HPA-5 system, among mothers of newborns with thrombocytopenia in our population would allow the identification of pregnancies at risk of alloimmune thrombocytopenia.

Adolescent↗
PubMed2006

Junctional adhesion molecule-C regulates vascular endothelial permeability by modulating VE-cadherin-mediated cell-cell contacts.

We recently reported that junctional adhesion molecule (JAM)-C plays a role in leukocyte transendothelial migration. Here, the role of JAM-C in vascular permeability was investigated in vitro and in vivo. As opposed to macrovascular endothelial cells that constitutively expressed JAM-C in cell-cell contacts, in quiescent microvascular endothelial cells, JAM-C localized mainly intracellularly, and was recruited to junctions upon short-term stimulation with vascular endothelial growth factor (VEGF) or histamine. Strikingly, disruption of JAM-C function decreased basal permeability and prevented the VEGF- and histamine-induced increases in human dermal microvascular endothelial cell permeability in vitro and skin permeability in mice. Permeability increases are essential in angiogenesis, and JAM-C blockade reduced hyperpermeability and neovascularization in hypoxia-induced retinal angiogenesis in mice. The underlying mechanisms of the JAM-C-mediated increase in endothelial permeability were studied. JAM-C was essential for the regulation of endothelial actomyosin, as revealed by decreased F-actin, reduced myosin light chain phosphorylation, and actin stress fiber formation due to JAM-C knockdown. Moreover, the loss of JAM-C expression resulted in stabilization of VE-cadherin-mediated interendothelial adhesion in a manner dependent on the small GTPase Rap1. Together, through modulation of endothelial contractility and VE-cadherin-mediated adhesion, JAM-C helps to regulate vascular permeability and pathologic angiogenesis.

Animals↗
PubMed2006

A variable number of tandem repeats polymorphism influences the transcriptional activity of the neonatal Fc receptor alpha-chain promoter.

The neonatal Fc receptor, FcRn, plays a central role in immunoglobulin G (IgG) transport across placental barriers. Genetic variations of FcRn-dependent transport across the placenta may influence antibody-mediated pathologies of the fetus and the newborn. Sequencing analysis of 20 unrelated individuals demonstrated no missense mutation within the five exons of the FcRn gene. However, a variable number of tandem repeats (VNTR) region within the FcRn promoter was observed, consisting of five different alleles (VNTR1-VNTR5). Alleles with two (VNTR2) and three (VNTR3) repeats were found to be most common in Caucasians (7.5 and 92.0%, respectively). Real-time polymerase chain reaction revealed that monocytes from VNTR3 homozygous individuals express 1.66-fold more FcRn transcript than do monocytes from VNTR2/VNTR3 heterozygous individuals (P = 0.002). In reporter plasmid assays, the VNTR3 allele supported the transcription of a reporter gene twice as effectively as did the VNTR2 allele (P = 0.003). Finally, under acidic conditions, monocytes from VNTR3 homozygous individuals showed an increased binding to polyvalent human IgG when compared with monocytes from VNTR2/VNTR3 heterozygous individuals (P = 0.021). These data indicate that a VNTR promoter polymorphism influences the expression of the FcRn receptor, leading to different IgG-binding capacities.

Alleles↗
PubMed2006

A naturally occurring LeuVal mutation in beta3-integrin impairs the HPA-1a epitope: the third allele of HPA-1.

BACKGROUND: Single-amino-acid substitution Leu33Pro in the beta3-integrin is responsible for the formation of the human platelet antigen (HPA)-1. Alloimmunization against HPA-1a (beta3-Leu33) is the most frequent cause of neonatal alloimmune thrombocytopenia and posttransfusion purpura. STUDY DESIGN AND METHODS: While HPA-1 genotyping a large cohort of patients with thromboembolic disease with a thermal cycler (LightCycler), one patient was identified with a unique HPA-1a melting curve. RESULTS: Sequence analysis revealed a C-to-G transversion at nucleotide 175 in the beta3-integrin (ITGB3) gene that alters the Leu33 codon to Val33. Further genotyping of healthy blood donors (n = 2950) identified one nonrelated Pro33Val33-positive individual. To examine whether the presence of Val33 affected the binding pattern of HPA-1 alloantibodies, transfectants were generated expressing recombinant beta3-Leu33 or beta3-Val33. Interestingly, differences in the reactivity of anti-HPA-1a were observed, with some HPA-1a alloantibodies showing diminished reactivity with beta3-Val33 compared to beta3-Leu33 and others reacting equally with both types. Similar findings were observed with recombinant human HPA-1a antibodies, with one of the three not binding to beta3-Val33. CONCLUSIONS: Our results demonstrate that the naturally occurring Leu33Val mutation in the beta3-integrin can disrupt some HPA-1a epitopes. These findings provide evidence for a heterogeneous humoral response against HPA-1a that may have potential clinical implications for alloimmune thrombocytopenia disorders.

Adult↗
PubMed2006

Human platelets target dendritic cell differentiation and production of proinflammatory cytokines.

BACKGROUND: Dendritic cells (DCs) are the most potent antigen-presenting cells that initiate and regulate immune responses. They are unique in their feature to produce bioactive interleukin (IL)-12, a major proinflammatory cytokine connecting innate and adaptive immunity. Platelets (PLTs) are highly reactive components of the circulatory system with fundamental importance in hemostasis and innate immunity. Recently, immunomodulatory capacities of single specific human PLT-derived products on DC effector functions were identified. To improve the understanding of PLT-DC interactions, this study investigates the influence of intact resting and activated PLTs on DC phenotype and key functions. STUDY DESIGN AND METHODS: Magnetic beads sorted CD14+ cells were expanded in the presence and absence of resting or activated PLTs. DC differentiation, maturation, allostimulatority capacity, antigen uptake, and cytokine profile were estimated to control group. RESULTS: Activated PLTs potently impaired DC differentiation according to CD1a expression (mean reduction, 62%; p < 0.05). Production of IL-12p70 and tumor necrosis factor-alpha was reduced in the presence of resting (mean reduction, 46 and 55%, respectively; p < 0.05) as well as activated PLTs (mean reduction, 63 and 49%, respectively; p < 0.05). In contrast to the suppression of proinflammatory cytokines, activated PLTs increased production of the immunoregulatory cytokine IL-10 by DCs (mean increase, 52%; p < 0.05). DC allostimulatority capacity, antigen uptake, and phenotypic maturation remained unaffected. CONCLUSION: It is proposed that intact PLTs connect immunity and hemostasis by interfering with DC differentiation and cytokine production. This interference might be of importance in clinical settings, such as DC therapy and PLT transfusions.

Blood Platelets↗
PubMed2005

The role of junctional adhesion molecule-C (JAM-C) in oxidized LDL-mediated leukocyte recruitment.

The junctional adhesion molecule-C (JAM-C) was recently shown to be a counter receptor for the leukocyte beta2-integrin Mac-1 (CD11b/CD18), thereby mediating interactions between vascular cells, particularly in inflammatory cell recruitment. Here, we investigated the role of JAM-C in oxidized low-density lipoprotein (LDL)-mediated leukocyte recruitment. As compared with normal arteries, immunostaining of atherosclerotic vessels revealed a high expression of JAM-C in association with neointimal smooth muscle cells and the endothelium. Moreover, JAM-C was strongly up-regulated in the spontaneous early lesions in ApoE -/- mice. In vitro, cultured human arterial smooth muscle cells (HASMC) were found to express JAM-C, and oxLDL, as well as enzymatically modified LDL (eLDL) significantly up-regulated JAM-C on both HASMC and endothelial cells in a time- and dose-dependent manner. Although under quiescent conditions, JAM-C predominantly localized to interendothelial cell-cell contacts in close proximity to zonula occludens-1 (ZO-1), oxLDL treatment induced a disorganization of JAM-C localization that was no more restricted to the interendothelial junctions. JAM-C thereby mediated both leukocyte adhesion and leukocyte transendothelial migration upon oxLDL treatment of endothelial cells, whereas JAM-C on quiescent endothelial cells only mediates leukocyte transmigration. Thus, upon oxLDL stimulation endothelial JAM-C functions as both an adhesion, as well as a transmigration receptor for leukocytes. Taken together, JAM-C is up-regulated by oxLDL and may thereby contribute to increased inflammatory cell recruitment during atherosclerosis. JAM-C may therefore provide a novel molecular target for antagonizing interactions between vascular cells in atherosclerosis.

Aged↗
PubMed2005

The homophilic binding of junctional adhesion molecule-C mediates tumor cell-endothelial cell interactions.

The junctional adhesion molecule C (JAM-C) was recently shown to undergo a heterophilic interaction with the leukocyte beta2 integrin Mac-1, thereby mediating interactions between vascular cells in inflammatory cell recruitment. Here, the homophilic interaction of JAM-C is presented and functionally characterized to mediate tumor cell-endothelial cell interactions. Recombinant soluble JAM-C in fluid phase bound to immobilized JAM-C as assessed in a purified system; moreover, JAM-C-transfected Chinese hamster ovary (CHO) cells adhered to immobilized JAM-C. The homophilic interaction of JAM-C was mediated by the isolated amino-terminal Ig domain (D1), but not the carboxyl-terminal Ig domain (D2), of the molecule. Dimerization of JAM-A is dependent on the sequence RVE in the amino-terminal Ig domain. This motif is conserved in JAM-C (Arg64-Ile65-Glu66), and a single amino acid mutation in this motif (E66R) abolished the homophilic interaction of JAM-C. The lung carcinoma cell line NCI-H522 was found to express JAM-C. NCI-H522 cells adhered to immobilized JAM-C, as well as to JAM-C-transfected CHO cells, but not to mock-transfected CHO cells or to CHO cells transfected with the JAM-C mutant (E66R). Adhesion of NCI-H522 cells to JAM-C protein or JAM-C-transfected CHO cells was abolished in the presence of soluble JAM-C or the isolated D1. Furthermore, the adhesion of NCI-H522 cells to endothelial cells was significantly blocked by soluble JAM-C or the isolated D1. Thus, JAM-C undergoes a homophilic interaction via the Arg64-Ile65-Glu66 motif on the membrane-distal Ig domain of the molecule. The homophilic interaction of JAM-C can mediate tumor cell-endothelial cell interactions and may thereby be involved in the process of tumor cell metastasis.

Amino Acid Motifs↗
PubMed2005

Junctional adhesion molecules (JAM)-B and -C contribute to leukocyte extravasation to the skin and mediate cutaneous inflammation.

Leukocyte extravasation is a finely tuned process, in which transmigration is the final step. Transmigration depends on molecules located at borders of endothelial cells; e.g., junctional adhesion molecules (JAM-A, -B and -C). In vivo blockade of JAM-A lead to decreased migration of monocytes into the skin. In contrast, the role of JAM-B and -C in development of cutaneous inflammation is unknown. We therefore elicited an allergic contact dermatitis in mice using 2,4-dinitro-1-fluorobenzene. RT-PCR and immunofluorescent staining of healthy skin revealed a constitutive JAM-B (66.4%+/-6.7% of all vessels) and -C expression (88.6+/-13.2%), which remained constant after induction of contact dermatitis. Functional studies, in which either JAM-B or -C neutralizing antibodies were injected into sensitized mice prior to allergen challenge showed a concentration-dependent reduction of the contact dermatitis. Decreased ear swelling was accompanied by reduction of leukocyte infiltration as analyzed by hematoxylin and eosin (H&E) histology and enzyme activity. Combined antibody treatment at doses of 1.25 mg per kg bodyweight lead to additive inhibition of allergic contact dermatitis, indicating that JAM-B and -C may have distinct functions. In conclusion, interactions with JAM-B and -C are essential for development of cutaneous inflammation.

Animals↗
PubMed2005

Immunization against a low-frequency human platelet alloantigen in fetal alloimmune thrombocytopenia is not a single event: characterization by the combined use of reference DNA and novel allele-specific cell lines expressing recombinant antigens.

BACKGROUND: Fetal and neonatal alloimmune thrombocytopenia (FNAIT) is caused by maternal immunization against a fetal platelet (PLT) alloantigen. In cases of FNAIT attributed to low-frequency PLT alloantigens, the laboratory diagnosis is often hampered by the lack of adequate PLTs. STUDY DESIGN AND METHODS: Three families with maternal immunization against fetal PLT antigens were analyzed. In Family 1, previous immunization of another female or woman has been observed. In Families 2 and 3, newborns presented with the typical clinical picture of FNAIT. Genotyping was performed by polymerase chain reaction-restriction fragment length polymorphism and direct sequencing with reference to DNA from Epstein-Barr virus-transformed B-lymphoblastoid cell lines. Antibodies were characterized by glycoprotein (GP)-specific immunoassay with a panel of stable Chinese hamster ovary cell lines expressing low-frequency alloantigens. RESULTS: In three families, maternal immunization associated with the low-frequency alloantigens human PLT antigen (HPA)-8bw (Sra), HPA-11bw (Groa), and HPA-13bw (Sita) was identified. Maternal serum samples showed positive reactions in an antigen capture assay with cell lines carrying recombinant GP IIb/IIIa (HPA-8bw and -11bw) or GPIa/IIa (HPA-13bw), respectively. These results could be confirmed by genotyping analysis of fathers and newborns. CONCLUSION: This study demonstrates that cases of FNAIT attributed to low-frequency PLT alloantigens cannot be regarded as single events. The availability of reference DNA and cell lines expressing recombinant PLT alloantigens can facilitate their identification.

Antigens, Human Platelet↗
PubMed2005

Relevance of the HPA-15 (Gov) polymorphism on CD109 in alloimmune thrombocytopenic syndromes.

BACKGROUND: Alloantibodies against the human platelet (PLT) alloantigen (HPA)-15 system residing on CD109 can cause fetal and neonatal alloimmune thrombocytopenia (FNAIT), posttransfusion purpura, and PLT transfusion refractoriness. The detection of antibodies against HPA-15, however, is hampered by the variable low expression and instability of the CD109 molecule during preparation and storage. STUDY DESIGN AND METHODS: This study analyzed the occurrence of HPA-15 alloantibodies in 1403 patients: 930 FNAIT and 473 polytransfused (PT) patients by modified monoclonal antibody specific immobilization of PLT antigens (MAIPA) assay with well-defined phenotyped PLTs. A DNA typing technique was developed to confirm the phenotypes of PLT donors. B-cell lines were established as sources of reference DNA. RESULTS: Genotyping of 407 unrelated blood donors revealed the gene frequencies 0.512 and 0.488 for HPA-15a and -15b, respectively. Based on the selection of PLTs expressing high amounts of CD109 on the surface (mean fluorescence intensity ratio 4-5 on expression peak on Days 2-4 after apheresis) antibody screening by the MAIPA assay was performed. In total, 16 (1.1%) HPA-15 alloantibodies were found comprising four anti-HPA-15a and 12 anti-HPA-15b. Anti-HPA-15b without other PLT-reactive antibodies were detectable in three serum samples of PT patients. The incidence of HPA-15 alloimmunization in PT patients was significantly higher than in mothers with FNAIT (3.0% vs. 0.22%). In relation to all detected HPA-specific antibodies, HPA-15 is responsible for 6.2 percent of alloimmunizations. CONCLUSION: These observations indicate that alloimmunization against HPA-15 should be considered as a cause for immune thrombocytopenia, particularly in patients receiving multiple PLT transfusions.

Antibodies, Monoclonal↗
PubMed2005

Functional heterogeneity of alloantibodies against the human platelet antigen (HPA)-1a.

The integrin alphaIIbbeta is the major fibrinogen receptor on the platelet membrane and plays a crucial role for platelet aggregation. The beta3-subunit carries the human platelet alloantigen (HPA)-1a, which is the main target for alloantibodies (alloabs) responsible for foetal and neonatal alloimmune thrombocytopenia (FNAIT) and post-transfusion purpura (PTP). Whereas PTP is almost invariably associated with severe bleeding, the clinical presentation of FNAIT ranges from mild thrombocytopenia to severe haemorrhagic diathesis. However, this clinical heterogeneity is not fully understood as it is not explained solely by the variability of the platelet count. Here, we examined the ability of HPA-1a alloabs from mothers with FNAIT (n = 43) and PTP patients (n = 8) to inhibit cell adhesion to fibrinogen and asked if this inhibition was correlated with the heterogeneity of the clinical picture. Stably transfected cells expressing HPA-1a (beta3-Leu33) and -1b (beta3-Pro33) isoforms were incubated with sera containing HPA-1a alloabs and were allowed to adhere to immobilised fibrinogen. The inhibitory activity was measured as percentage of cell adhesion in the presence of patient sera versus normal AB serum. Only two FNAIT sera specifically inhibited the adhesion of HPA-1a, but not HPA-1b cells. Two other FNAIT sera blocked the adhesion of HPA-1a as well as HPA-1b cells. Interestingly, all four neonates with inhibitory HPA-1a alloabs (9% of all sera) suffered severe bleeding. In comparison, the majority of PTP sera (75%) inhibited cell binding to fibrinogen, four PTP sera selectively inhibited the adhesion of HPA-1a cells whilst 2 sera impaired the binding of both allotypes. Our observations indicate that 1) HPA-1a alloabs are heterogeneous in their ability to interfere with fibrinogen binding, and 2) inhibition of the alphaIIbbeta3 fibrinogen receptor by HPA-1a alloabs may contribute to pronounced bleeding in patients with alloimmune thrombocytopenia.

Animals↗
PubMed2004

The junctional adhesion molecule-C promotes neutrophil transendothelial migration in vitro and in vivo.

The third member of the family of junctional adhesion molecules (JAMs), JAM-3, also called JAM-C, was recently shown to be a novel counter-receptor on platelets for the leukocyte beta(2)-integrin Mac-1 (alphaMbeta(2), CD11b/CD18). Here, new functional aspects of the role of endothelial cell JAM-C were investigated. Endothelial cells express JAM-C, which is predominantly localized within junctions at interendothelial contacts, since it codistributes with a tight junction component, zonula occludens-1. Whereas JAM-C does not participate in neutrophil adhesion to endothelial cells, it mediates neutrophil transmigration in a Mac-1-dependent manner. In particular, inhibition of JAM-C significantly reduced neutrophil transendothelial migration, and the combination of JAM-C and platelet/endothelial cell adhesion molecule-1 blockade almost completely abolished neutrophil transendothelial migration in vitro. In vivo, inhibition of JAM-C with soluble mouse JAM-C resulted in a 50% reduction of neutrophil emigration in the mouse model of acute thioglycollate-induced peritonitis. Thus, JAM-C participates in neutrophil transmigration and thereby provides a novel molecular target for antagonizing interactions between vascular cells that promote inflammatory vascular pathologies.

Animals↗
PubMed2004

Human alloantibody anti-Mart interferes with Mac-1-dependent leukocyte adhesion.

The CD11b/CD18 integrin plays a crucial role in cell-cell adhesion processes. Recently, we described a case of severe neonatal alloimmune neutropenia (NAIN) caused by an alloantibody against a variant of the CD11b subunit (Mart alloantigen). Allele-specific transfected cells allowed us to demonstrate that an H61R point mutation is directly responsible for the formation of Mart epitopes. No difference in the adhesion capability between H61 and R61 homozygous neutrophils was observed. Functional analysis showed that anti-Mart inhibited Mac-1-dependent adhesion of neutrophils and monocytic U937 cells to fibrinogen, intercellular adhesion molecule-1 (ICAM-1), receptor for advanced glycation end product (RAGE), and glycoprotein Ibalpha but not to junctional adhesion molecule-C or urokinase plasminogen activator receptor (uPAR). Accordingly, anti-Mart blocked neutrophil and U937 cell adhesion to endothelial cells and platelet-leukocyte aggregate formation in whole blood under high shear. Other sera of anti-Mart from mothers of infants without NAIN did not show inhibitory properties. We conclude that anti-Mart antibodies with different functional properties exist. This is supported by our findings that anti-Mart antibodies have different abilities to inhibit cell-cell adhesion, to enhance the respiratory burst of neutrophils, and to recognize different epitopes at the N-terminal region of CD11b. In conclusion, some anti-Mart alloantibodies interfere with Mac-1-dependent cellular functions of neutrophils, cause NAIN, and may be used as tools for studying Mac-1-dependent functions.

Animals↗
PubMed2004

Ceftriaxone causes drug-induced immune thrombocytopenia and hemolytic anemia: characterization of targets on platelets and red blood cells.

BACKGROUND: Ceftriaxone, a third-generation cephalosporin, has been reported to occasionally cause fatal drug-induced immune hemolytic anemia (DIHA). A clinical and serologic analysis of the first two patients with severe drug-induced thrombocytopenia (DITP) due to ceftriaxone and one patient with fatal DIHA is reported. STUDY DESIGN AND METHODS: Sera were assessed by the IAT, EIA, glycoprotein (GP)-specific immunoassay, flow cytometry, and immunoprecipitation using transfectants expressing GPIIb/IIIa and GPIb/IX and with different cephalosporins. RESULTS: Sera from Patients 1 and 2 reacted strongly with PLTs in the presence of the drug, but not with RBCs. The binding sites of the drug-dependent antibodies (DDAbs) could be localized to GPIIb/IIIa and GPIb/IX, respectively. Inhibition studies indicated that DDAbs recognized epitopes residing on the GPIIb/IIIa complex and on the GPIX subunit, respectively. No cross-reactivity was observed with other cephalosporin derivatives. Serum 3 showed strong agglutination with RBCs of Rh(null) phenotype in the presence of ex-vivo metabolites of ceftriaxone, but no cross-reactivity with PLTs. CONCLUSIONS: The first two cases of severe DITP and a third patient with DIHA are reported. DDAbs from all patients showed individual reaction patterns and clear cell lineage specificity. In addition, the DDAbs were dependent on the substitution at position 3 of the ceftriaxone molecule. Epitopes on GPIIb/IIIa and GPIX were involved on PLTs. The Rh protein was not the only target of DDAbs on RBCs.

Adolescent↗
PubMed2004

Diclofenac-induced antibodies against red blood cells are heterogeneous and recognize different epitopes.

BACKGROUND: Diclofenac (DCF) is a widely used nonsteroidal anti-inflammatory drug implicated as a cause of immune hemolytic anemia. Drug derivatives have been suggested to be an important-or probably the primary-immunizing agent in drug-induced immune reactions. A systematic evaluation of 12 patients with DCF-induced immune hemolysis is reported. STUDY DESIGN AND METHODS: All sera samples were evaluated with standard serologic tests for the detection of red blood cells (RBCs) and platelet (PLT) antibodies in the presence of DCF, DCF-urine (DCF-U), and five chemically defined metabolites. RESULTS: Twelve patients' sera samples reacted with DCF-U, but only 9 reacted with DCF. When derivatives were tested, no metabolite was recognized by all sera samples (although 4'-OH-DCF was recognized by 11/12), and no metabolite remained unrecognized. As demonstrated with Rh(null) cells, the Rh complex may represent an important, but not the only, target protein for which drug-dependent antibodies are specific. PLT-reactive antibodies were not detectable. CONCLUSION: There is evidence that patients with DCF-induced immune hemolysis produce a broad spectrum of anti-DCF/RBC antibodies. 4'-OH-DCF seems to represent the most immunogenic metabolite. Nevertheless, all patients' sera samples contain a mixture of antibodies that recognize several and distinguishable epitopes. These epitopes consist of different drug metabolites and a target protein on the RBC surface, which appears to be the Rh complex in many, but not in all, cases. Additional target proteins remain to be identified.

Anemia, Hemolytic, Autoimmune↗
PubMed2004

Glanzmann thrombasthenia Frankfurt I is associated with a point mutation Thr176Ile in the N-terminal region of alpha IIb subunit integrin.

In this study, we report on the characterization of a patient with Glanzmann thrombasthenia (GT). Immunochemical analysis on platelets from the patient showed that the expression of alpha IIb beta 3 was only 25% of that in normal healthy controls, suggesting a case of GT. Functional analysis revealed a total lack of fibrinogen binding capacity. Molecular genetic analysis of the full-length cDNA sequences of alpha IIb and beta 3 subunits showed a novel point mutation C621T in alpha IIb cDNA, leading to a missense substitution of threonine for isoleucine at position 176. Coexpression of normal beta 3 and mutant alpha IIb(1176) isoform in mammalian cells showed a marked reduction in the expression of alpha IIb beta 3 heterodimer when compared to the wild-type and a decreased intracellular level of alpha IIb. The T176 I mutation is located in the N-terminal region in the W3:1-2 connecting strand of the beta-propeller. These data suggest that the N-terminal alpha IIb domain plays an important structural role in the formation of heterodimer and that it is also involved in fibrinogen binding.

Blood Coagulation Tests↗
PubMed2003

High molecular weight kininogen regulates platelet-leukocyte interactions by bridging Mac-1 and glycoprotein Ib.

Leukocyte-platelet interaction is important in mediating leukocyte adhesion to a thrombus and leukocyte recruitment to a site of vascular injury. This interaction is mediated at least in part by the beta2-integrin Mac-1 (CD11b/CD18) and its counter-receptor on platelets, glycoprotein Ibalpha (GPIbalpha). High molecular weight kininogen (HK) was previously shown to interact with both GPIbalpha and Mac-1 through its domains 3 and 5, respectively. In this study we investigated the ability of HK to interfere with the leukocyte-platelet interaction. In a purified system, HK binding to GPIbalpha was inhibited by HK domain 3 and the monoclonal antibody (mAb) SZ2, directed against the epitope 269-282 of GPIbalpha, whereas mAb AP1, directed to the region 201-268 of GPIbalpha had no effect. In contrast, mAb AP1 inhibited the Mac-1-GPIbalpha interaction. Binding of GPIbalpha to Mac-1 was enhanced 2-fold by HK. This effect of HK was abrogated in the presence of HK domains 3 or 5 or peptides from the 475-497 region of the carboxyl terminus of domain 5 as well as in the presence of mAb SZ2 but not mAb AP1. Whereas no difference in the affinity of the Mac-1-GPIbalpha interaction was observed in the absence or presence of HK, maximal binding of GPIbalpha to Mac-1 doubled in the presence of HK. Moreover, HK/HKa increased the Mac-1-dependent adhesion of myelomonocytic U937 cells and K562 cells transfected with Mac-1 to immobilized GPIbalpha or to GPIbalpha-transfected Chinese hamster ovary cells. Finally, Mac-1-dependent adhesion of neutrophils to surface-adherent platelets was enhanced by HK. Thus, HK can bridge leukocytes with platelets by interacting via its domain 3 with GPIbalpha and via its domain 5 with Mac-1 thereby augmenting the Mac-1-GPIbalpha interaction. These distinct molecular interactions of HK with leukocytes and platelets contribute to the regulation of the adhesive behavior of vascular cells and provide novel molecular targets for reducing atherothrombotic pathologies.

Animals↗