Sesamin's Therapeutic Actions on Cyclophosphamide-Induced Hepatotoxicity, Molecular Mechanisms, and Histopathological Characteristics.

Cyclophosphamide, an alkylating agent integral to specific cancer chemotherapy protocols, is often curtailed in application owing to its significant hepatotoxic side effects. Therefore, this study was conducted to assess the hepatoprotective potential of sesamin, a plant-originated antioxidant, using rat models. The rats were divided into five groups: a control group received only the vehicle for six days; a cyclophosphamide group received an intraperitoneal (i.p.) single injection of cyclophosphamide (150 mg/kg) on day four; a sesamin group received a daily high oral dose (20 mg/kg) of sesamin for six days; and two groups were pretreated with oral sesamin (10 and 20 mg/kg daily from day one to day six) followed by an i.p. injection of cyclophosphamide on day four. The final and last sesamin dose was administered 24 h before euthanasia. At the end of the experiment, blood and liver tissue were collected for biochemical and histopathological assessments. The results indicated significantly increased liver markers (AST, ALT, ALP, and BIL), cytokines (TNFα and IL-1ß), caspase-3, and malondialdehyde (MDA) in the cyclophosphamide group as compared to the normal control. Additionally, there was a significant decline in antioxidants (GSH) and antioxidant enzymes (CAT and SOD), but the sesamin treatment reduced liver marker enzymes, cytokines, and caspase-3 and improved antioxidants and antioxidant enzymes. Thus, sesamin effectively countered these alterations and helped to normalize the histopathological alterations. In conclusion, sesamin demonstrated the potential for attenuating cyclophosphamide-induced hepatotoxicity by modulating cytokine networks, apoptotic pathways, and oxidative stress, suggesting its potential role as an adjunct in chemotherapy to reduce hepatotoxicity.

Thymoquinone Ameliorates Carfilzomib-Induced Renal Impairment by Modulating Oxidative Stress Markers, Inflammatory/Apoptotic Mediators, and Augmenting Nrf2 in Rats.

Chemotherapy-induced kidney damage is an emerging problem that restricts cancer treatment effectiveness. The proteasome inhibitor carfilzomib (CFZ) is primarily used to treat multiple myeloma and has been associated with severe renal injury in humans. CFZ-induced nephrotoxicity remains an unmet medical need, and there is an urgent need to find and develop a nephroprotective and antioxidant therapy for this condition. Thymoquinone (TQ) is a bioactive compound that has been isolated from Nigella sativa seeds. It has a wide range of pharmacological properties. Therefore, this experimental design aimed to study the effectiveness of TQ against CFZ-induced renal toxicity in rats. The first group of rats was a normal control (CNT); the second group received CFZ (4 mg/kg b.w.); the third and fourth groups received TQ (10 and 20 mg/kg b.w.) 2 h before receiving CFZ; the fifth group received only TQ (20 mg/kg b.w.). This experiment was conducted for 16 days, and at the end of the experiment, blood samples and kidney tissue were collected for biochemical assays. The results indicated that administration of CFZ significantly enhanced serum marker levels such as BUN, creatinine, and uric acid in the CFZ group. Similarly, it was also noticed that CFZ administration induced oxidative stress by reducing antioxidants (GSH) and antioxidant enzymes (CAT and SOD) and increasing lipid peroxidation. CFZ treatment also enhanced the expression of IL-1ß, IL-6, and TNF-α production. Moreover, CFZ increased caspase-3 concentrations and reduced Nrf2 expression in the CFZ-administered group. However, treatment with 10 and 20 mg/kg TQ significantly decreased serum markers and increased antioxidant enzymes. TQ treatment considerably reduced IL-1ß, IL-6, TNF-α, and caspase-3 concentrations. Overall, this biochemical estimation was also supported by histopathological outcomes. This study revealed that TQ administration significantly mitigated the negative effects of CFZ treatment on Nrf2 expression. Thus, it indicates that TQ may have utility as a potential drug to prevent CFZ-induced nephrotoxicity in the future.

The Association between Constitutional Knee Alignment and the Horizontal Orientation of the Ankle Joint Line.

PURPOSE OF THE STUDY To characterize constitutional frontal alignment of the ankle in genua vara, valga, and norma. MATERIAL AND METHODS Long-leg standing radiographs of 589 patients presenting between 2011 and 2020 for knee-complaints because of any reason were chosen from our database. Cases with fractures or history of bony-realignment-surgeries were excluded. The Hip-Knee-Ankle angle (HKA), the mechanical Lateral Distal Tibia Angle (mLDTA), and the Tibia-Plafond-Horizontal-Orientation angle (TPHA) were measured in 354 patients. For this study, neutral frontal alignment of the leg was defined as HKA between -3.0° and +4.0°. HKA-values <-3.0° were defined as genua valga and values >4.0° were defined as genua vara. According to these cutoffs, data was categorized into the following three patient groups: genua vara (n=157), genua norma (n=106), genua valga (n=91). For each group, the ankle alignment in the frontal plane was compared to the HKA. Finally, the three groups were compared to each other. RESULTS In the varus-group, the HKA-value was 6.9°±2.4°, the TPHA-value was 4.7°±3.5°, and the mLDTA-value was 87.4°±4.8°. In the neutral-group, the HKA-value was 1.8°±2.0°, the TPHA-value was 2.5°±2.9°, and the mLDTA-value was 87.2°±4.6°. In the valgus-group, the HKA-value was -6.0°±2.7°, the TPHA-value was -0.2°±4.7°, and the mLDTA-value was 85.0°±4.7°. DISCUSSION The frontal alignment of the ankle joint line depends on the overall frontal alignment of the leg. The TPHA correlates with varus or valgus alignment of the knee, but the mLDTA does not. In patients with valgus-aligned long-leg axis, the TPHA demonstrated less valgus alignment than in patients with varus-aligned long-leg axis. This knowledge is especially useful when planning osteotomies for correction of lower extremity malalignment. CONCLUSIONS During the planning process of osteotomies around the knee, the TPHA should be appreciated because it correlates with the constitutional knee alignment. Key words: valgus, varus, frontal alignment, coronal alignment, osteotomy.

Tissue engineering approaches for the repair and regeneration of the anterior cruciate ligament: towards 3D bioprinted ACL-on-chip.

The anterior cruciate ligament (ACL) is the most frequently injured ligament in the knee. The current method to treat the injured ligament is reconstruction using autografts and allografts. Reconstruction requires the regeneration of ligament, bone and their interface to ensure proper recovery. Recently, researchers have focused on using tissue-engineered scaffolds made of synthetic materials and biomaterials -such as collagen, decellularised tissues, silk and synthetic polymers produced following different manufacturing methods - for ACL reconstruction,. Different materials can be easily processed using various fabrication methods for mimicking the mechanical properties of the ACL. The advances in technologies play an important role in the production of constructions that can mimic native ACL.. The present review addresses integrative scaffold design, different challenges in the potential materials and manufacturing methods as well as future strategies for ACL repair. Furthermore, the review provides a road map to 3D printing combined with organ-on-chip technology to demonstrate the potential for cost-effective and user-friendly fabrication methods for ACL engineering. Finally, it underlines the potential of 3D bioprinting and organ-on-chip technologies for micro-engineering of ligaments and their associated environment.

Molecular evidence and hematological alterations associated with the occurrence of coronavirus in domestic dogs in Pakistan.

Canine Enteric Coronavirus (CCoV) is one of the major enteric pathogen affecting dogs. This study aims to investigate the molecular prevalence, phylogenetic analysis, associated risk factors, and haemato-biochemical alterations in Canine Coronavirus in dogs in district Lahore, Pakistan. 450 fecal samples were collected from symptomatic dogs originating from various pet-clinics and kennels during 2018-2019. Samples were initially analyzed by sandwich lateral flow immunochromatographic assay and then further processed by RT-PCR (reverse transcriptase polymerase chain reaction) targeting the M gene followed by sequencing. RT-PCR based positive (n=20) and negative (n=20) dogs were samples for their blood for the haemato-biochemical analysis. A questionnaire was used to collect data from pet owners, in order to analyze the data for risk factors analysis by chi square test on SPSS. The prevalence of CCoV was 35.1%, and 23.8 % through Sandwich lateral flow immunochromatographic and RT-PCR respectively. Various risk factors like breed, age, sex, vomiting, diarrhea, sample source, body size, cohabitation with other animals, living environment, food, deworming history, contact with other animals or birds feces, and season were significantly associated with CCoV. The CCoV identified in Pakistan were 98% similar with the isolates from China (KT 192675, 1), South Korea (HM 130573, 1), Brazil (GU 300134, 1), Colombia (MH 717721, 1), United Kingdom (JX 082356, 1) and Tunisia (KX156806). Haematobiochemical alterations in CCoV affected dogs revealed anaemia, leucopenia, lymphopenia, neutrophilia, and decreased packed cell volume, and a significant increase in alkaline phosphate and alanine transaminase. It is concluded that infection with canine coronavirus appears widespread among dog populations in district Lahore, Pakistan. This study is the first report regarding the molecular detection and sequence analysis of CCoV in Pakistan.

Molecular evidence and hematological alterations associated with the occurrence of coronavirus in domestic dogs in Pakistan

@#Canine Enteric Coronavirus (CCoV) is one of the major enteric pathogen affecting dogs. This study aims to investigate the molecular prevalence, phylogenetic analysis, associated risk factors, and haemato-biochemical alterations in Canine Coronavirus in dogs in district Lahore, Pakistan. 450 fecal samples were collected from symptomatic dogs originating from various pet-clinics and kennels during 2018-2019. Samples were initially analyzed by sandwich lateral flow immunochromatographic assay and then further processed by RT-PCR (reverse transcriptase polymerase chain reaction) targeting the M gene followed by sequencing. RT-PCR based positive (n=20) and negative (n=20) dogs were samples for their blood for the haemato-biochemical analysis. A questionnaire was used to collect data from pet owners, in order to analyze the data for risk factors analysis by chi square test on SPSS. The prevalence of CCoV was 35.1%, and 23.8 % through Sandwich lateral flow immunochromatographic and RT-PCR respectively. Various risk factors like breed, age, sex, vomiting, diarrhea, sample source, body size, cohabitation with other animals, living environment, food, deworming history, contact with other animals or birds feces, and season were significantly associated with CCoV. The CCoV identified in Pakistan were 98% similar with the isolates from China (KT 192675, 1), South Korea (HM 130573, 1), Brazil (GU 300134, 1), Colombia (MH 717721, 1), United Kingdom (JX 082356, 1) and Tunisia (KX156806). Haematobiochemical alterations in CCoV affected dogs revealed anaemia, leucopenia, lymphopenia, neutrophilia, and decreased packed cell volume, and a significant increase in alkaline phosphate and alanine transaminase. It is concluded that infection with canine coronavirus appears widespread among dog populations in district Lahore, Pakistan. This study is the first report regarding the molecular detection and sequence analysis of CCoV in Pakistan.

Gunshot induced injuries in orthopaedic trauma research. A bibliometric analysis of the most influential literature.

A growing burden of gunshot injuries demands evidence-based ballistic trauma management. No comprehensive systematic overview of the current knowledge is available to date. This study aims to identify and analyze the most influential publications in the field of orthopedic ballistic trauma research. All databases available in the Thomson Reuters Web of Knowledge were searched to conduct this bibliometrical study. The most cited orthopedic ballistic trauma articles published between 1950 and 2015 were identified by use of a multi-step approach. Publications with ten citations and more were analyzed for citations, journal, authorship, geographic origin, area of research, anatomical site, study type, study category, and level of evidence. Citations of the 128 included studies ranged from 113 to 10. These were published in fifty different journals between 1953 and 2011. Most publications (n=106; 83%) originated from the USA, were retrospective (n=85; 66.4%), level IV studies (n=90; 70.3%), reported on spinal gunshot injuries (n=49; 38.33%) and were published between 1980 and 2000 (n=111; 86.7%). This bibliometric study provides the first comprehensive overview of influential publications in the field of orthopedic ballistic trauma research. More prospective studies and high-quality systematic reviews are needed. Centres with a high burden of gunshot injuries from the developing world need to share their experience in form of international publications, to provide a more comprehensive picture of the global gun-related orthopedic injury burden. TYPE OF STUDY: bibliometric analysis: level III.

Dynamic intraligamentary stabilisation: initial experience with treatment of acute ACL ruptures.

AIMS: The purpose of this study was to report the experience of dynamic intraligamentary stabilisation (DIS) using the Ligamys device for the treatment of acute ruptures of the anterior cruciate ligament (ACL). PATIENTS AND METHODS: Between March 2011 and April 2012, 50 patients (34 men and 16 women) with an acute rupture of the ACL underwent primary repair using this device. The mean age of the patients was 30 years (18 to 50). Patients were evaluated for laxity, stability, range of movement (ROM), Tegner, Lysholm, International Knee Documentation Committee (IKDC) and visual analogue scale (VAS) scores over a follow-up period of two years. RESULTS: At final follow-up, anteroposterior translation differed from the normal knee by a mean of 0.96 mm (-2 mm to 6 mm). Median (interquartile range) IKDC, Tegner, Lysholm and VAS scores were 98 (95 to 100), 6 (5 to 7), 100 (98 to 100) and 10 (9 to 10), respectively. Pre-injury Tegner activity levels were reached one year post-operatively. A total of nine patients (18%) required a secondary intervention; five developed instability, of whom four underwent secondary hamstring reconstructive surgery, and five required arthroscopic treatment for intra-articular impingement due to scar tissue which caused a fixed flexion deformity. In addition, 30 patients (60%) required removal of the tibial screw. CONCLUSION: While there was a high rate of secondary interventions, 45 patients (90%) retained their repaired ACL two years post-operatively, with good clinical scores and stability of the knee. TAKE HOME MESSAGE: Dynamic intraligamentary stabilisation presents a promising treatment option for acute ACL ruptures, eliminating the need for ACL reconstruction. Cite this article: Bone Joint J 2016;98-B:793-8.

Sero-prevalence of Hepatitis B and C Virus from rural areas of northern Punjab (Sargodha District), Pakistan.

Pakistan is endemic for hepatitis B virus (HBV) and hepatitis C virus (HCV) infections with 10 and 11 million infections, respectively. The epidemiological studies of these virus showed that the information is only from few cities of the country and is relevant to high risk groups. It is of great importance to have an idea about the prevalence of infectious agents in general population to help in identification of hot spot for infections. Identification of hot spots will help in disease management for future. As the there is no report form district Sargodha (Punjab Province) so this study was designed to analyze the prevalence of HBV and HCV in general population. Blood samples of 2373 randomly selected individuals from six different tehsils were collected and were analyzed for HBV and HCV sero-positivity. An overall prevalence of both HBV and HCV in district Sargodha was 28.10% (667/2373). HCV prevalence was (20.01%) and HBV seropositivity was (8.09%). Males were more infected than females, and a significant difference was found in positive cases between male (58.77%) and female (41.22%). The most common routes of transmission of hepatitis virus in present study were shaving assisted by barbers in male patients 143 (21.43%), non sterile or used needles and syringes 127 (19.04%), dental surgical procedures 88 (13.19%), and sharing razors in males 49 (7.34%). In female patients a significant factor is labor and child birth process. Most cases of hepatitis were seen in mesons, farmers and house wives. The prevalence of HBV and HCV in general population of district Sargodha is very high. The study will help for better management of disease to contain the disease spread. The study highlighted that District Sargodha is endemic for these viral infections and it is highly warranted to carry out more studies to get better idea about the infection spread. Community education campaigns are also highly warranted to general population as well as high risk population to control future disease spread.

Mapping tibiofemoral gonarthrosis: an MRI analysis of non-traumatic knee cartilage defects.

OBJECTIVE: Arthroscopy is "the gold standard" for the diagnosis of knee cartilage lesions. However, it is invasive and expensive, and displays all the potential complications of an open surgical procedure. Ultra-high-field MRI now offers good opportunities for the indirect assessment of the integrity and structural changes of joint cartilage of the knee. The goal of the present study is to determine the site of early cartilaginous lesions in adults with non-traumatic knee pain. METHODS: 3-T MRI examinations of 200 asymptomatic knees with standard and three-dimensional double-echo steady-state (3D-DESS) cartilage-specific sequences were prospectively studied for early degenerative lesions of the tibiofemoral joint. Lesions were classified and mapped using the modified Outerbridge and modified International Cartilage Repair Society classifications. RESULTS: A total of 1437 lesions were detected: 56.1% grade I, 33.5% grade II, 7.2% grade III and 3.3% grade IV. Cartographically, grade I lesions were most common in the anteromedial tibial areas; grade II lesions in the anteromedial L5 femoral areas; and grade III in the centromedial M2 femoral areas. CONCLUSION: 3-T MRI with standard and 3D-DESS cartilage-specific sequences demonstrated that areas predisposed to early osteoarthritis are the central, lateral and ventromedial tibial plateau, as well as the central and medial femoral condyle. ADVANCES IN KNOWLEDGE: In contrast with previous studies reporting early cartilaginous lesions in the medial tibial compartment and/or in the medial femoral condyle, this study demonstrates that, regardless of grade, lesions preferentially occur at the L5 and M4 tibial and L5 and L2 femoral areas of the knee joint.

Primary testicular lymphoma.

Primary testicular non-Hodgkin lymphoma (PTL) comprises around 9% of testicular cancers and 1-2% of all non-Hodgkin lymphomas. Its incidence is increasing and it primarily affects older men, with a median age at presentation of around 67 years. By far the most common histological subtype is diffuse large B-cell lymphoma, accounting for 80-90% of PTLs. Most patients present with a unilateral testicular mass or swelling. Up to 90% of patients have stage I or II disease at diagnosis (60 and 30%, respectively) and bilateral testicular involvement is seen in around 35% of patients. PTL demonstrates a continuous pattern of relapse and propensity for extra-nodal sites such as the central nervous system and contralateral testis. Retrospective data have emphasised the importance of prophylactic radiotherapy in reducing recurrence rates within the contralateral testis. Recent outcome data from the prospective IELSG-10 trial have shown far better progression-free and overall survival than historical outcomes. This supports the use of orchidectomy followed by Rituximab- cyclophosphamide, doxorubicin, vincristine and prednisolone (R-CHOP), central nervous system prophylaxis and prophylactic radiotherapy to the contralateral testis with or without nodal radiotherapy in patients with limited disease. Central nervous system relapse remains a significant issue and future research should focus on identifying the best strategy to reduce its occurrence. Here we discuss the evidence supporting combination chemotherapy and radiotherapy in PTL.

Vicinal thiols are required for activation of the αIIbß3 platelet integrin.

BACKGROUND: Closely spaced thiols in proteins that interconvert between the dithiol form and disulfide bonds are called vicinal thiols. These thiols provide a mechanism to regulate protein function. We previously found that thiols in both αIIb and ß3 of the αIIbß3 fibrinogen receptor were required for platelet aggregation. METHODS AND RESULTS: Using p-chloromercuribenzene sulfonate (pCMBS) we provide evidence that surface thiols in αIIbß3 are exposed during platelet activation. Phenylarsine oxide (PAO), a reagent that binds vicinal thiols, inhibits platelet aggregation and labeling of sulfhydryls in both αIIb and ß3. For the aggregation and labeling studies, binding of PAO to vicinal thiols was confirmed by reversal of PAO binding with the dithiol reagent 2,3-Dimercapto-1-propanesulfonic acid (DMPS). In contrast, the monothiol ß-mercaptoethanol did not reverse the effects of PAO. Additionally, PAO did not inhibit sulfhydryl labeling of the monothiol protein albumin, confirming the specificity of PAO for vicinal thiols in αIIbß3. As vicinal thiols represent redox sensitive sites that can be regulated by reducing equivalents from the extracellular or cytoplasmic environment, they are likely to be important in regulating activation of αIIbß3. Additionally, when the labeled integrin was passed though a lectin column containing wheat germ agglutinin and lentil lectin a substantial amount of non-labeled αIIbß3 eluted separately from the labeled receptor. This suggests that two populations of integrin exist on platelets that can be distinguished by thiol labeling. CONCLUSION: A vicinal thiol-containing population of αIIbß3 provides redox sensitive sites for regulation of αIIbß3.

Acute unilateral cataract in a postpartum adolescent with poorly-controlled type 1 diabetes.

Acute cataract is recognized as a rare complication in adolescents with type 1 diabetes mellitus and may be associated with rapid improvement in glycaemia in patients with newly diagnosed diabetes. Transient cataracts, which resolve following improved metabolic control, and irreversible cataracts requiring surgery have also previously been documented. Development or progression of retinopathy may complicate pregnancy in women with diabetes. To our knowledge, we present the first case report of an acute cataract developing postpartum in a woman with type 1 diabetes. This rare case serves to demonstrate a possible association between acute cataract and altered glycaemic control in pregnancy. Acute cataract should be considered in any woman with diabetes who develops sudden visual loss following pregnancy.

Patient attitudes towards medical students in Riyadh, Saudi Arabia.

CONTEXT: A key element of medical education is patients' willingness to and cooperation in involving a medical student in their care. OBJECTIVE: To assess patients' attitudes and their associated factors toward involving medical students in their care. METHODS: This study was conducted over a period of two months in two hospitals in Riyadh, Saudi Arabia. Six hundred patients were approached to participate in outpatient and inpatient settings of four medical specialty services. A pre-tested questionnaire was used to collect the data. RESULTS: There were 492 respondents (82%), of whom 50% were males, 74% were married, and 51% were outpatients. Sixty percent identified medical students on the team of doctors that had treated them. Eighty-one percent of respondents felt that students' general appearance and manner were important to their willingness to have them participate in their care, and 64% would not object to the presence of medical students during physical examinations. Most (63%) preferred to be informed about students' involvement beforehand. Fifty-five percent believed that they had the right to refuse medical students, with women, married and less educated individuals and those seen in particular departments more likely to believe so. The preferred role for students also varied with gender, marital status and department. CONCLUSION: This study finds that most Saudi patients view medical students' involvement in their care positively. They prefer to be told in advance about students' presence, asked for their permission, and have students' involvement restricted to certain times.

Cemento-ossifying fibroma--a rare case.

The concept of 'fibro-osseous lesions' of bone has evolved over the last several decades and now includes two major entities: fibrous dysplasia and ossifying fibroma, as well as the other less common lesions such as florid osseous dysplasia, periapical dysplasia, focal sclerosing osteomyelitis, proliferative periostitis of garrie, and ostitis deformans. The cemento-ossifying fibroma is a central neoplasm of bone as well as periodontium which has caused considerable controversy because of confusion regarding terminology and the criteria for its diagnosis. The cemento-ossifying fibroma is odontogenic in origin, whereas ossifying fibroma is of bony origin. This article reports a rare case of an 11-year-old male who came to us with the history of swelling in the maxillary anterior region causing difficulty in closing of mouth as well as in mastication.

A practical profile of integrated safety assessment of near-surface disposal of radwaste at PINSTECH.

Near-surface or shallow land disposal of radioactive waste has been the primary practice at the Pakistan Institute of Nuclear Science and Technology (PINSTECH). The adopted choice of this mode of disposal has been based on a study of the site and the quality and quantity of waste generated at the 5 MW reactor with HEU fuel. Specific measures regarding the radiation safety of the workers and environmental protection have been adopted. The waste disposal operations are conducted to meet local regulatory requirements, IAEA recommendations and internationally endorsed principles such as ALARA (as low as reasonably achievable - economic, social and other relevant factors being considered). The data obtained through the years of operational and management experience have manifested the robustness of the disposal system and reliability of the disposal criterion, and have also served to further refine the latter. Consequently, confidence in the current shallow-land-burial practices has increased. Radiological safety of these practices has been assessed by addressing different aspects of the safety and disposal system. These parameters, as indices of a non-exclusive and operational safety model, are presented.

Masses of nasal cavity, paranasal sinuses and nasopharynx: A clinicopathological study.

An analysis of clinicopathological features of 240 cases presenting as mass in nasal cavity, paranasal sinuses and nasopharynx observed, both retrospectively and prospectively, over a period of 5 years in Jawaharlal Nehru Medical College. Aligarh. The incidence of masses in nasal cavity, paranasal sinuses and nasopharynx was 34.3 cases per year. Amongst the 240 cases studied, there were 144 cases (60%) of non-neoplastic lesions, 56 cases (23.33%) of benign lesions and 40 cases (16.67%) of malignant lesions. All age groups were involved and the mean age of presentation with the increasing age were: - non-neoplastic (22.5 years), benign tumors (26.8 years) and malignant tumors (35.3 years). The male to female ratio was 1.7:1 for non-neoplastic lesions; 3:1 for benign tumors: and 2.3:1 for malignant lesions. In this study maximum number of cases were present in nasal cavity (65%) followed by paranasal sinuses (20%) and least number of cases involved the nasopharynx (15%). The relative number of non-neoplastic and neoplastic lesions varies from region to region. A provisional diagnosis was made after clinical assessment and radiological investigation but final diagnosis was made after histopathological examination.

Binding studies of the enzyme (factor IXa) with the cofactor (factor VIIIa) in the assembly of factor-X activating complex on the activated platelet surface.

Activated platelet membranes expose binding sites for the enzyme factor (F)IXa, the substrate (FX) and the cofactor (FVIIIa) that colocalize to assemble the FX-activating complex and promote optimal rates of FX activation. To determine the stoichiometry and affinity of binding to activated platelets, coordinate, equilibrium binding studies with enzyme (125I-FIXa) and cofactor (131I-FVIII or 131I-FVIIIa) were carried out in the presence of saturating concentrations of substrate (FX). Results of these studies indicate that in the presence of FX (1.5 micro m), the enzyme (active-site-inhibited Glu-Gly-Arg-FIXa, EGR-FIXa) and procofactor (FVIII) bind to an equal number (approximately 700 sites/platelet) of receptors whereas the active cofactor (FVIIIa) binds an additional approximately 500 high-affinity FVIIIa binding sites per platelet (Kd approximately 0.8 nm). With excess zymogen (FIX) to block shared FIX/FIXa-binding sites, the stoichiometry of 125I-FIXa and 131I-FVIIIa binding was 1:4. These FIXa/FVIIIa binding studies together with previously reported evidence of the coordinate binding of FVIIIa and FX to equivalent numbers of binding sites on activated platelets provide strong evidence to support the conclusion that FVIIIa comprises the receptor that presents FX to FIXa for efficient catalysis on the activated platelet membrane.

The assembly of the factor X-activating complex on activated human platelets.

Platelet membranes provide procoagulant surfaces for the assembly and expression of the factor X-activating complex and promote the proteolytic activation and assembly of the prothrombinase complex resulting in normal hemostasis. Recent studies from our laboratory and others indicate that platelets possess specific, high-affinity, saturable, receptors for factors XI, XIa, IX, IXa, X, VIII, VIIIa, V, Va and Xa, prothrombin, and thrombin. Studies described in this review support the hypothesis that the factor X-activating complex on the platelet surface consists of three receptors (for the enzyme, factor IXa; the substrate, factor X; and the cofactor, factor VIIIa), the colocalization of which results in a 24 million-fold acceleration of the rate of factor X activation. Whether the procoagulant surface of platelets is defined exclusively by procoagulant phospholipids, or whether specific protein receptors exist for the coagulant factors and proteases, is currently unresolved. The interaction between coagulation proteins and platelets is critical to the maintenance of normal hemostasis and is pathogenetically important in human disease.