Microbial Fibrinolytic Enzymes as Anti-Thrombotics: Production, Characterisation and Prodigious Biopharmaceutical Applications

Cardiac disorders such as acute myocardial infarction, embolism and stroke are primarily attributed to excessive fibrin accumulation in the blood vessels, usually consequential in thrombosis. Numerous methodologies including the use of anti-coagulants, anti-platelet drugs, surgical operations and fibrinolytic enzymes are employed for the dissolution of fibrin clots and hence ameliorate thrombosis. Microbial fibrinolytic enzymes have attracted much more attention in the management of cardiovascular disorders than typical anti-thrombotic strategies because of the undesirable after-effects and high expense of the latter. Fibrinolytic enzymes such as plasminogen activators and plasmin-like proteins hydrolyse thrombi with high efficacy with no significant after-effects and can be cost effectively produced on a large scale with a short generation time. However, the hunt for novel fibrinolytic enzymes necessitates complex purification stages, physiochemical and structural-functional attributes, which provide an insight into their mechanism of action. Besides, strain improvement and molecular technologies such as cloning, overexpression and the construction of genetically modified strains for the enhanced production of fibrinolytic enzymes significantly improve their thrombolytic potential. In addition, the unconventional applicability of some fibrinolytic enzymes paves their way for protein hydrolysis in addition to fibrin/thrombi, blood pressure regulation, anti-microbials, detergent additives for blood stain removal, preventing dental caries, anti-inflammatory and mucolytic expectorant agents. Therefore, this review article encompasses the production, biochemical/structure-function properties, thrombolytic potential and other surplus applications of microbial fibrinolytic enzymes.

Vascular injury in macroscopically normal skin of patients with severe COVID-19 infection: clinical-pathologic correlations

Objectives: Taking into account that the documentation of the histopathological features in severe disease caused by SARS-CoV-2 has been scarce due to the avoidance of performing autopsies, the aim of the study was to detect the microscopic changes associated with severe COVID-19 infection in normal-appearing skin, without prominent dermatologic signs of a generalized microvascular thrombotic disorder, in accordance with the clinical evolution of disease. Methods: In this morphological and immunohistochemical study we included cutaneous biopsy samples from 12 symptomatic patients with severe and critical type SARS-CoV-2 infection (with the admission date between February and June 2020), treated in the Intensive Therapy Unit Care of Emergency County Hospital Targu-Mures, Romania. Results: The average age of our patients was 65.18 ± 14.21 years (range 41 to 83), and 66.67% of the patients were male. The histological and immunohistochemical assessment of cutaneous biopsies: in 4 cases the histological examination revealed small fibrin thrombi in deep-seated venules and small veins of subcuticular adipose tissue, and also 4 cutaneous biopsies showed occlusive vascular thrombosis in association with massive perivascular inflammatory infiltrate destroying and compromising the integrity of the vessel wall. The immunohistochemical examination of the composition of perivascular inflammatory infiltrate showed a predominance of CD3 positive lymphocytes, admixed with CD68 positive Mo/ MF, some of them activated with FXIII expression. In the perivascular infiltrate, the presence of granulocytes and B lymphocytes was not characteristic. Conclusion: According to our observations, in severe COVID-19, the cutaneous tissue is involved even in the absence of clinically obvious changes. Due to the relatively easy accessibility of skin samples, these could be applied to determine the severity of the patient’s clinical status, and to predict the necessity for anti-complement or anticoagulant treatments in the early stages of a severe SARS-CoV-2 infection.

Angiolipoma in a Dog

Background: Angiolipoma is a benign tumor composed of endothelial cells and mature adipocytes. Tumors reported in domestic species include two variants; infiltrative or non-infiltrative. Bitches and intact males seem predisposed. This mesenchyme tumor is commonly mistaken with lipoma due to its soft texture and subcutaneous site and often requires histopathology to confirm its diagnosis. Microscopic examination also enables the evaluation of surgical margins and rule out possible infiltrative sites. Complete surgical excision is usually curative. This study reports a case of non-infiltrating angiolipoma in a dog.Case: A 14-year-old mixed-breed dog was presented to a veterinary clinic in the city of Rio de Janeiro, RJ, Brazil. On palpation, a painless mass was noted, with high mobility and covered by intact hirsute skin in the right subcutaneous ventrolateral region. Computed tomography of the chest showed an expansive mass of uptake only from the edges of the soft tissues of the right subcutaneous ventrolateral region. The mass was homogeneous and well delimited, suggesting a neoplastic process. Subsequently, the mass was surgically removed, fixed in 10% buffered formalin, and sent for histopathological analysis. On macroscopic examination, the mass was well delimited, without skin coverage, and measured 2.3 × 1.9 × 0.6 cm. The consistency was smooth and unctuous in appearance with a compact cream-colored surface with blackish multifocal spots. Under microscopy, the histological sections showed neoplasm of mature adipocytes and of endothelial cells of blood vessels benign were filled with a marked amount of red blood cells. Multifocal fibrin thrombi and a mild inflammatory infiltrate composed of lymphocytes and rare mast cells were evident. There was no infiltration in the regional skeletal musculature. Thus, a diagnosis of non-infiltrative angiolipoma was established.Discussion: The diagnosis of non-infiltrating angiolipoma in this case was established through the results of histopathological examination. The occurrence of this neoplasm in dogs is uncommon, and the data reported in the veterinary medicine literature are scarce. However, in this study, it was found that the neoplasm on screening presented a behavior like that of lipomas, with noninvasive growth and the absence of local recurrence. The canine species does not commonly convey pain on palpation during a clinical examination, as observed in the present case. In humans, multiple angiolipoma nodules are common; this clinical presentation differs from that in animals, in which solitary nodules are generally observed. In dogs, as in the present case, they seem to have a predilection for the trunk. In animals, the pathogenesis of angiolipomas is not established, but in humans, it is based on theories that include the reaction to harmful stimuli and congenital malformation of adipose tissue. In humans, the presence of fibrin thrombi on the periphery of the region of cell proliferation are microscopic findings that can assist in the diagnosis of angiolipomas, an approach that was implemented in the present case. The occurrence of this neoplasm in dogs is uncommon, and the data reported in the veterinary medicine literature are scarce. The importance of an adequate description of angiolipomas is based on the need to provide information about its epidemiology, biological behavior, and prognosis.

Pfeifer-Weber-Christian Disease and Benign Multiple Subcutaneous Noninfiltrative Angiolipomas: A Puzzling Case and Review of a Rare Entitety

Background: Pfeifer-Weber-Christian disease (PWCD) is one of many rare diseases that may be easily missed if there is not a high degree of suspicion. Angiolipomas are rare, benign subcutaneous tumors, composed of adipose tissue and blood vessels and often containing fibrin thrombi. The majority of angiolipomas occur sporadically; however, there is a minority of cases that have been associated with long-term corticosteroid use. Case presentation: We report here an unusual case of PWCD associated with benign multiple subcutaneous noninfiltrative angiolipomas confirmed by skin biopsy. Systemic corticosteroid therapy was not effective at reducing flares of panniculitis, and during this therapy angiolipomas gradually increased in size. In contrast, administration of oral Cyclosporine A (CyA) led to a rapid remission of the PWCD. Conclusions: As PWCD has no known aetiology and no specific treatment has been established, the successful therapy with the CyA supports the hypothesis that PWCD is a T cell mediated autoinflammatory condition. Also, this case represent very rare side effects of corticosteroid therapy, such as induction of de novo angiolipomas or increased growth of existing tumours. This case bring diagnostic difficulties in everyday clinical practice, especially in patients with panniculitides, but histopathological evaluation usually resolves the dilemma.

Necrotising Vasculitis in Covid-19: watch out for bowel perforation

Introduction: SARS-Cov-2 infection can be asymptomatic, greatly impair respiratory function and, sometimes, affect other organs. Gastro-enteric involvement seems to be not so rare and many patients suffered for abdominal pain, anorexia, nausea and vomiting, diarrhoea or jaundice. Case presentation: Here we report a case of a Covid-19 patient who developed a right colon perforation due to a pseudomembranous colitis without Clostridium Difficile toxins finding associated to a necrotising vasculitis. In this patient a PCR test performed on gastric fluid showed SARS-CoV-2 enteric replication. Discussion: Necrotizing vasculitis is an anatomopathological typical findings in Covid-19. It has been documented in several organs and tissues. In our case, evident foci of necrotizing vasculitis with intravascular obliteration by fibrin thrombi with macrophagic cells infiltration are anatomopathological findings of a CD toxins free pseudomembranous colitis leading to bowel perforation. Conclusions: Bowel perforation due to necrotizing vasculitis leading to pseudomembranous colitis could be a SARS-Cov-2 related clinical finding.

The Spectrum of Histopathologic Findings in Lungs of Patients With Fatal Coronavirus Disease 2019 (COVID-19) Infection

Context.— Respiratory failure appears to be the ultimate mechanism of death in most patients with severe coronavirus disease 2019 (COVID-19) infection. Studies of postmortem COVID-19 lungs largely report diffuse alveolar damage and capillary fibrin thrombi, but we have also observed other patterns. Objective.— To report demographic and radiographic features along with macroscopic, microscopic, and microbiologic postmortem lung findings in patients with COVID-19 infections. Design.— Patients with confirmed COVID-19 infection and postmortem examination (March 2020–May 2020) were included. Clinical findings were abstracted from medical records. Lungs were microscopically reviewed independently by 4 thoracic pathologists. Imaging studies were reviewed by a thoracic radiologist. Results.— Eight patients (7 men, 87.5%; median age, 79 years; range, 69–96 years) died within a median of 17 days (range, 6–100 days) from onset of symptoms. The median lung weight was 1220 g (range, 960–1760 g); consolidations were found in 5 patients (62.5%) and gross thromboemboli were noted in 1 patient (12.5%). Histologically, all patients had acute bronchopneumonia; 6 patients (75%) also had diffuse alveolar damage. Two patients (25%) had aspiration pneumonia in addition. Thromboemboli, usually scattered and rare, were identified in 5 patients (62.5%) in small vessels and in 2 of these patients also in pulmonary arteries. Four patients (50%) had perivascular chronic inflammation. Postmortem bacterial lung cultures were positive in 4 patients (50%). Imaging studies (available in 4 patients) were typical (n = 2, 50%), indeterminate (n = 1, 25%), or negative (n = 1, 25%) for COVID-19 infection. Conclusions.— Our study shows that patients infected with COVID-19 not only have diffuse alveolar damage but also commonly have acute bronchopneumonia and aspiration pneumonia. These findings are important for management of these patients.

Pulmonary post-mortem findings in a large series of COVID-19 cases from Northern Italy

ABSTRACTImportanceThe analysis of lung tissues of patients with COVID-19 may help understand pathogenesis and clinical outcomes in this life-threatening respiratory illness.ObjectiveTo determine the histological patterns in lung tissue of patients with severe COVID-19.Design and participantsLungs tissues of 38 cases who died for COVID-19 in two hospital of Northern Italy were systematically analysed. Hematoxylin-eosin staining, immunohistochemistry for the inflammatory infiltrate and cellular components, electron microscopy were performed.ResultsThe features of the exudative and proliferative phases of Diffuse Alveolar Disease (DAD) were found: capillary congestion, necrosis of pneumocytes, hyaline membrane, interstitial oedema, pneumocyte hyperplasia and reactive atypia, platelet-fibrin thrombi. The inflammatory infiltrate was composed by macrophages in alveolar lumens and lymphocytes mainly in the interstice. Electron microscopy revealed viral particles in the cytoplasm of pneumocytes.Conclusions and relevanceThe predominant pattern of lung lesions in COVID-19 patients is DAD, as described for the other two coronavirus that infect humans, SARS-CoV and MERS-CoV. Hyaline membrane formation and pneumocyte atypical hyperplasia are frequently found. The main relevant finding is the presence of platelet-fibrin thrombi in small arterial vessels; this important observation fits into the clinical context of coagulopathy which dominates in these patients and which is one of the main targets of therapy.

Intravascular Papillary Endothelial Hyperplasia With a Phlebolith of the Tongue: A Potential Pitfall

Intravascular papillary endothelial hyperplasia (IPEH) or so-called Masson hemangioma is a benign nonneoplastic thrombus-associated organizing proliferation. De novo or secondary IPEHs are not uncommon oral lesions. An associated oral phlebolith is uncommon. Oral IPEHs and phleboliths can be diagnostically challenging. Certain histomorphologic features are helpful clues. Immunohistochemistry is a useful confirmatory tool. We present a case of an IPEH with phlebolith in a 37-year-old healthy lady who presented with a nodular lesion of the tongue. Histology showed a calcific nodule associated with an endothelial-lined vascular lesion with histomorphologic and immunohistochemical features of IPEH. The presence of a solitary extruded phlebolith could be challenging for pathologists and could be confused with other oral calculi, particularly sialoliths. The presence of papillary fronds, endothelial-lined minute vascular channels, fibrin thrombi, and hemosiderin pigment on the surface of the calculus points to a calcified thrombus. The lamellated concentric onion ring layers corresponding to Zahn lines around a central calcific nidus is another hint. Phleboliths should be considered by pathologists in their differential diagnosis of oral calculi. Distinction from sialolith is clinically important because the etiology and management are different.