Fibrinogenolysis and fibrinolysis in vaccine-induced immune thrombocytopenia and thrombosis.

BACKGROUND: Vaccine-induced immune thrombocytopenia and thrombosis (VITT) is a rare syndrome associated with adenoviral vector vaccines for COVID-19. The syndrome is characterized by thrombosis, anti-platelet factor 4 (PF4) antibodies, thrombocytopenia, high D-dimer, and hypofibrinogenemia. OBJECTIVES: To investigate abnormalities in fibrinolysis that contribute to the clinical features of VITT. METHODS: Plasma samples from 18 suspected VITT cases were tested for anti-PF4 by ELISA and characterized as meeting criteria for VITT (11/18) or deemed unlikely (7/18; non-VITT). Antigen levels of PAI-1, factor XIII (FXIII), plasmin-α2antiplasmin (PAP), and inflammatory markers were quantified. Plasmin generation was quantified by chromogenic substrate. Western blotting was performed with antibodies to fibrinogen, FXIII-A, and plasminogen. RESULTS: VITT patients 10/11 had scores indicative of overt disseminated intravascular coagulation, while 0/7 non-VITT patients met the criteria. VITT patients had significantly higher levels of inflammatory markers, IL-1ß, IL-6, IL-8, TNFα, and C-reactive protein. In VITT patients, both fibrinogen and FXIII levels were significantly lower, while PAP and tPA-mediated plasmin generation were higher compared to non-VITT patients. Evidence of fibrinogenolysis was observed in 9/11 VITT patients but not in non-VITT patients or healthy controls. Fibrinogen degradation products were apparent, with obvious cleavage of the fibrinogen α-chain. PAP complex was evident in those VITT patients with fibrinogenolysis, but not in non-VITT patients or healthy donors. CONCLUSION: VITT patients show evidence of overt disseminated intravascular coagulation and fibrinogenolysis, mediated by dysregulated plasmin generation, as evidenced by increased PAP and plasmin generation. These observations are consistent with the clinical presentation of both thrombosis and bleeding in VITT.

Venous thromboembolism risk following temporary immobilisation after injury: evaluation of the Aberdeen VTE risk tool.

BACKGROUND: Temporary lower limb immobilisation following injury is a risk factor for symptomatic venous thromboembolism (VTE). Pharmacological thromboprophylaxis can mitigate this risk but it is unclear which patients benefit from this intervention. The Aberdeen VTE risk tool was developed to tailor thromboprophylaxis decisions in these patients and this evaluation aimed to describe its performance in clinical practice. Secondarily, diagnostic metrics were compared with other risk assessment methods (RAMs). METHODS: A prospective cohort service evaluation was conducted. Adult patients (≥16 years) managed with lower limb immobilisation for injury who were evaluated with the Aberdeen VTE risk tool prior to discharge from the ED were identified contemporaneously between February 2014 and December 2020. Electronic patient records were scrutinised up to 3 months after removal of immobilisation for the development of symptomatic VTE or sudden death due to pulmonary embolism (PE). Other RAMs, including the Thrombosis Risk Prediction for Patients with cast immobilisation (TRiP(cast)) and Plymouth scores, were assimilated retrospectively and diagnostic performance compared. RESULTS: Of 1763 patients (mean age 46 (SD 18) years, 51% women), 15 (0.85%, 95% CI 0.52% to 1.40%) suffered a symptomatic VTE or death due to PE. The Aberdeen VTE tool identified 1053 (59.7%) patients for thromboprophylaxis with a sensitivity of 80.0% (95% CI 54.8% to 93.0%) and specificity of 40.4% (95% CI 38.1% to 42.6%) for the primary outcome. In 1695 patients, fewer were identified as high risk by the TRiP(cast) (33.3%) and Plymouth (24.4%) scores, but with greater specificity, 67.0% and 75.6%, respectively, than dichotomous RAMs, including the Aberdeen VTE tool. CONCLUSION: Routine use of the Aberdeen VTE tool in our population resulted in an incidence of symptomatic VTE of less than 1%. Ordinal RAMs, such as the TRiP(cast) score, may more accurately reflect VTE risk and permit more individually tailored thromboprophylaxis decisions but prospective comparison is needed.

Skin cancer in essential thrombocythaemia and polycythaemia vera patients treated with hydroxycarbamide.

Hydroxycarbamide (HC) is used as a cytoreductive treatment in myeloproliferative neoplasms (MPN). Observational studies have raised the possibility that HC contributes to the development of secondary malignancies, including skin tumours in MPN patients. In this retrospective observational study, we report a single-centre experience of 324 HC-treated MPN patients with long-term follow-up, compared to 47 MPN patients not on HC. Thirty-three patients (10.2%) (HC) versus one patient (2.1%) (no HC) developed skin tumours during follow-up (Hazard ratios [HR] 5.70, 95% confidence intervals 0.66-48.09, p = 0.112). However, male gender, age at MPN diagnosis, type of MPN (polycythaemia rubra vera) and previous history of skin cancer were prognostic variables associated with development of skin cancer.

Spinal cord compression secondary to extramedullary haematopoiesis in transformed polycythaemia rubra vera.

Polycythaemia vera (PV) is a clonal proliferative disorder of the bone marrow characterised by autonomous haematopoiesis, which results in a panmyelosis in the peripheral blood. It is typically characterised by an acquired mutation in JAK-2 V617F. Progression to myelofibrosis (MF), characterised by worsening cytopenias and the development of constitutional symptoms, is seen in up to 10% of cases. Extramedullary haematopoiesis (EMH) in the spleen is a common finding in myelofibrotic transformation, but elsewhere in the body it is extremely unusual. We report the case of a 69-year-old male whose PV progressed to secondary MF and who presented with compression of the thoracic spinal cord directly as a result of EMH. Cytogenetic and molecular findings in the bone marrow were in keeping with evolving myeloid disease. He was managed by surgical laminectomy with an excellent outcome. Extramedullary haematopoiesis may be seen in both PV and on transformation to MF. This very rare complication should be borne in mind when managing patients with myeloproliferative disorders.

The suboptimal fibrinolytic response in COVID-19 is dictated by high PAI-1.

BACKGROUND: Severe COVID-19 disease is associated with thrombotic complications and extensive fibrin deposition. This study investigates whether the hemostatic complications in COVID-19 disease arise due to dysregulation of the fibrinolytic system. METHODS: This prospective study analyzed fibrinolytic profiles of 113 patients hospitalized with COVID-19 disease with 24 patients with non-COVID-19 respiratory infection and healthy controls. Antigens were quantified by Ella system or ELISA, clot lysis by turbidimetric assay, and plasminogen activator inhibitor-1 (PAI-1)/plasmin activity using chromogenic substrates. Clot structure was visualized by confocal microscopy. RESULTS: PAI-1 and its cofactor, vitronectin, are significantly elevated in patients with COVID-19 disease compared with those with non-COVID-19 respiratory infection and healthy control groups. Thrombin activatable fibrinolysis inhibitor and tissue plasminogen activator were elevated in patients with COVID-19 disease relative to healthy controls. PAI-1 and tissue plasminogen activator (tPA) were associated with more severe COVID-19 disease severity. Clots formed from COVID-19 plasma demonstrate an altered fibrin network, with attenuated fiber length and increased branching. Functional studies reveal that plasmin generation and clot lysis were markedly attenuated in COVID-19 disease, while PAI-1 activity was elevated. Clot lysis time significantly correlated with PAI-1 levels. Stratification of COVID-19 samples according to PAI-1 levels reveals significantly faster lysis when using the PAI-1 resistant (tPA) variant, tenecteplase, over alteplase lysis. CONCLUSION: This study shows that the suboptimal fibrinolytic response in COVID-19 disease is directly attributable to elevated levels of PAI-1, which attenuate plasmin generation. These data highlight the important prognostic potential of PAI-1 and the possibility of using pre-existing drugs, such as tenecteplase, to treat COVID-19 disease and potentially other respiratory diseases.

Luminal Bioavailability of Orally Administered ω-3 PUFAs in the Distal Small Intestine, and Associated Changes to the Ileal Microbiome, in Humans with a Temporary Ileostomy.

BACKGROUND: Oral administration of purified omega-3 (ω-3) PUFAs is associated with changes to the fecal microbiome. However, it is not known whether this effect is associated with increased PUFA concentrations in the gut. OBJECTIVES: We investigated the luminal bioavailability of oral ω-3 PUFAs (daily dose 1 g EPA and 1g DHA free fatty acid equivalents as triglycerides in soft-gel capsules, twice daily) and changes to the gut microbiome, in the ileum. METHODS: Ileostomy fluid (IF) and blood were obtained at baseline, after first capsule dosing (median 2 h), and at a similar time after final dosing on day 28, in 11 individuals (median age 63 y) with a temporary ileostomy. Fatty acids were measured by LC-tandem MS. The ileal microbiome was characterized by 16S rRNA PCR and Illumina sequencing. RESULTS: There was a mean 6.0 ± 9.8-fold and 6.6 ± 9.6-fold increase in ileal EPA and DHA concentrations (primary outcome), respectively, at 28 d, which was associated with increased RBC ω-3 PUFA content (P ≤ 0.05). The first oral dose did not increase the ileal ω-3 PUFA concentration except in 4 individuals, who displayed high luminal EPA and DHA concentrations, which reduced to concentrations similar to the overall study population at day 28, suggesting physiological adaptation. Bacteroides, Clostridium, and Streptococcus were abundant bacterial genera in the ileum. Ileal microbiome variability over time and between individuals was large, with no consistent change associated with acute ω-3 PUFA dosing. However, high concentrations of EPA and DHA in IF on day 28 were associated with higher abundance of Bacteroides (r2 > 0.86, P < 0.05) and reduced abundance of other genera, including Actinomyces (r2 > 0.94, P < 0.05). CONCLUSIONS: Oral administration of ω-3 PUFAs leads to increased luminal ω-3 PUFA concentrations and changes to the microbiome, in the ileum of individuals with a temporary ileostomy. This study is registered on the ISRCTN registry as ISRCTN14530452.

Does age-adjusted D-Dimer have a role in assessment of VTE recurrence rates?

INTRODUCTION: It is known that D-dimer levels increase with age and several studies have evaluated the use of an age-adjusted (AA) cut-off in the initial assessment of suspected venous thromboembolism (VTE). We performed a retrospective study to assess the effect that using an AA D-dimer in the DASH score would have on the recommended duration of anticoagulant treatment for patients following a first unprovoked episode of VTE and then compared this with the advice that has been given to patients using a fixed cut-off D-dimer in the DASH score. METHODS: Data were collected for the period from April 2014 to October 2017. For each patient, the DASH score by a single cut-off D-dimer value (500 ng/mL) as well as an AA D-dimer cut-off value (D-dimer cut-off value equal to age in years × 10) was calculated for patients over 50 years. The Vienna prediction model was employed alongside this to compare the VTE recurrence risk using a well-established method. RESULTS: A total of 204 patients were eligible for analysis, 145 of whom were over the age of 50 years. In 24 of these patients, the use of the AA D-dimer made a significant impact on the predicted risk of recurrence using the DASH score and would have likely changed the recommendation to offer long-term anticoagulation. CONCLUSION: As an age-adjusted D-dimer cut-off has been assessed in the diagnostic setting, it would be logical and appropriate to also consider this philosophy in the prediction of the risk of recurrence of VTE following a first unprovoked event.

Limited hybridization between introduced and Critically Endangered indigenous tilapia fishes in northern Tanzania.

Hybridization between introduced and indigenous species can lead to loss of unique genetic resources and precipitate extinction. In Tanzania, the Nile tilapia (Oreochromis niloticus) and blue-spotted tilapia (Oreochromis leucostictus) have been widely introduced to non-native habitats for aquaculture and development of capture fisheries. Here, we aimed to quantify interspecific hybridization between these introduced species and the indigenous species Oreochromis esculentus, Oreochromis jipe and Oreochromis korogwe. In the Pangani basin, several hybrids were observed (O. niloticus × O. jipe, O. leucostictus × O. jipe, O. niloticus × O. korogwe), although hybrids were relatively uncommon within samples relative to purebreds. Hybrids between the native O. jipe × O. korogwe were also observed. In the Lake Victoria basin, no evidence of hybrids was found. Analysis of body shape using geometric morphometrics suggested that although purebreds could be discriminated from one another, hybrids could not be readily identified on body and head shape alone. These results provide the first evidence of hybridization between the introduced species and the Critically Endangered O. jipe in Tanzania. Given uncertainty regarding benefits of introduced species over large-bodied indigenous species in aquaculture and capture fisheries, we suggest that future introductions of hybridization-prone species should be carefully evaluated.

Reducing the use of inappropriate coagulation testing in emergency general surgical patients.

Background and aims Indiscriminate coagulation testing in emergency general surgical patients can lead to inappropriate delay in surgery, cause unnecessary concern and is associated with significant cost. The British Committee for Standards in Haematology recommends against coagulation testing to predict peri-operative bleeding risk in unselected patients. Our aim was to assess the appropriateness of coagulation tests performed in emergency general surgical patients and evaluate the effect of a series of educational interventions on clinical practice. Methods and results Appropriate indications for performing coagulation testing included a positive bleeding history, the presence of liver disease/cholestasis, sepsis or use of anticoagulants. Initial data on 142 patients were collected over 2 weeks of receiving. Following analysis, indications for appropriate coagulation testing were highlighted and data were collected on a further 190 patients. Comparing the audit cycles, we observed a decrease in the proportion of patients who underwent routine testing (49.3% vs 32.6%; p = 0.002) and inappropriate testing (67% of tests vs 34% of tests; p < 0.001). Despite being highlighted, there was no evidence of improved documentation of bleeding histories on admission. Conclusions This observational study suggests that simple educational messages can reduce the inappropriate use of coagulation screening tests in general surgical emergencies. This seems to result from clarification of the appropriate surgical indications for coagulation testing in this group.

A randomised trial of the effect of omega-3 polyunsaturated fatty acid supplements on the human intestinal microbiota.

OBJECTIVE: Omega-3 polyunsaturated fatty acids (PUFAs) have anticolorectal cancer (CRC) activity. The intestinal microbiota has been implicated in colorectal carcinogenesis. Dietary omega-3 PUFAs alter the mouse intestinal microbiome compatible with antineoplastic activity. Therefore, we investigated the effect of omega-3 PUFA supplements on the faecal microbiome in middle-aged, healthy volunteers (n=22). DESIGN: A randomised, open-label, cross-over trial of 8 weeks' treatment with 4 g mixed eicosapentaenoic acid/docosahexaenoic acid in two formulations (soft-gel capsules and Smartfish drinks), separated by a 12-week 'washout' period. Faecal samples were collected at five time-points for microbiome analysis by 16S ribosomal RNA PCR and Illumina MiSeq sequencing. Red blood cell (RBC) fatty acid analysis was performed by liquid chromatography tandem mass spectrometry. RESULTS: Both omega-3 PUFA formulations induced similar changes in RBC fatty acid content, except that drinks were associated with a larger, and more prolonged, decrease in omega-6 PUFA arachidonic acid than the capsule intervention (p=0.02). There were no significant changes in α or ß diversity, or phyla composition, associated with omega-3 PUFA supplementation. However, a reversible increased abundance of several genera, including Bifidobacterium, Roseburia and Lactobacillus was observed with one or both omega-3 PUFA interventions. Microbiome changes did not correlate with RBC omega-3 PUFA incorporation or development of omega-3 PUFA-induced diarrhoea. There were no treatment order effects. CONCLUSION: Omega-3 PUFA supplementation induces a reversible increase in several short-chain fatty acid-producing bacteria, independently of the method of administration. There is no simple relationship between the intestinal microbiome and systemic omega-3 PUFA exposure. TRIAL REGISTRATION NUMBER: ISRCTN18662143.

Defective α2 antiplasmin cross-linking and thrombus stability in a case of acquired factor XIII deficiency.

Acquired factor XIII (FXIII) deficiency is a rare and life-threatening condition that is often misdiagnosed or missed completely. A 72-year-old woman presented with symptoms of major unprovoked bleeding but routine coagulation screening tests and platelet count were normal. Low activated FXIII (FXIIIa) activity levels and abnormal urea clot stability led to diagnosis of acquired FXIII deficiency. A modified Bethesda inhibitor titre of 1.6 Bethesda units/ml indicated the presence of a FXIII inhibitor. Bleeding responded to a single dose of FXIII concentrate and immunosuppression with prednisolone induced remission. A subsequent relapse was treated with combined prednisolone and Rituximab resulting in a prolonged, ongoing remission. Here we analyse the mechanisms underlying this idiopathic case of acquired FXIII deficiency. Prospective analysis of patient plasma revealed minimal FXIIIa activity and antigen in presentation and relapse samples. Thrombi formed from these samples lysed rapidly and showed an absence of cross-linked α2 AP. Western blotting revealed the presence of FXIII-B, indicating only FXIII-A and FXIII-A2 B2 were affected. FXIII activity and antigen levels normalised on remission. Our data suggest the presence of inhibitor-induced clearance of FXIII from plasma. As a consequence, reduced thrombus stability was evident due to defective α2 AP cross-linking, thereby explaining symptoms of excessive bleeding.

Does a second resection provide a survival benefit in patients diagnosed with incidental T1b/T2 gallbladder cancer following cholecystectomy?

BACKGROUND: Incidental T1b/T2 gallbladder cancers are often managed with a second resection. However it is unclear whether the additional surgical risk is associated with any survival advantage. The aim of this study was to examine the outcomes of patients who underwent a second resection following a diagnosis of incidental T1b/T2 gallbladder cancer. METHODS: A retrospective analysis of patients undergoing surgical management following a diagnosis of incidental T1b/T2 gallbladder cancer between 1994 and 2014. Survival outcomes were analysed using the Kaplan-Meier method. RESULTS: Twenty two patients underwent completion surgery following diagnosis of T1b/T2 gallbladder cancer at initial cholecystectomy, 11 of which were found to have residual disease. The presence of residual disease at second surgery in T1b/T2 disease was associated with worse overall survival (residual disease: median survival 12 months, absence of residual disease: median survival not reached, p = 0.025). CONCLUSION: A significant percentage of patients with T1b/T2 disease have identifiable residual disease following second surgery. Residual disease is associated with poor survival. It is therefore important to inform patients that completion cholecystectomy is primarily performed to inform staging rather than to improve prognosis.