chuka_lis (chuka_lis) wrote,
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О патологических поражениях легких

Статья от шведов.
Что у них вышло, по результатам аутопсий: те, кто умер именно от ковидного ТОРС (без прочих "отягчающих"), почти не имели воздуха в легких, и легкие были отечные- пространство внутри альвеол заполнено, всем, чем можно, кроме возудха. Собственно тромбов было мало, а "закупорки", преимущественно, были в альвеолоах- иммунные и прочие. А те, что в сосудах- происходили из-за агрегации тромбоцитов. Сосуды возле пострадавших от вируса альвеол были тоже  дефектные- с "дырками" и пр.
В легких шли активные регенерационные и пролиферативные процессы- во всех тканях, чтобы восстановить обширные повреждения, нанесенные вирусом- именно эти процессы давали картину "консолидации".
Вирус не размножался в эндотелиальных клетках (внутри- не было)- но все они в пристуствии вируса (снаружи и в соседнихк летках) были поврежденными, и не могли функционировать адекватно.
Эпителий верхних дыхательных путей и бронхов  у большинства -не был поражен, и не имел вируса (не смотря на АСЕ рецепторы). В легких, коронавирус размножался в макрофагах и миелоидных клетках определенного типа, несущих ресепторы КД68, кроме того, что активно реплицировался в альвеолах- в пневмоцитах первого и второго типа. Более всего коронавируса было в альвеолах. Паренхима легких у тяжелых больных с ТОРС была поражена коронавирусом сильнее всего остального, особенно полны вируса были "гиалиновые мембраны" альвеол. Авторы выдвигают предположение, что, скорее всего, и выделяться коронавирус может, находясь внутри полуразрушенных клеток легких (а составе фрагментов гиалиновых мембран), вместе с ними, с "жидкостью" при выдыхании- что обеспечивает ему дольшую сохранность в окружающей среде, и попадание потом, при высыхании, в легкие другого человека, и далее- в легочные макрофаги (которые поглощают всякие подобные загрязнители). А в них вирус "высвобождается", после того, как клетка-носитель или капсула, разрушается ферментами макрофага, и вирус заражает таким образом макрофаг. Если не этот механизм- то авторы думают, что причиной обнаруженного ими инфицирования макрофагов может быть проникновение вируса посредством Fc- рецепторов (антительные), и тогда их инфицирование является результатом АЗУИ. Если это так, то это надо быстренько хорошо исследовать- иначе часть вакцинированных может получить серьезную болезнь (ТОРС) или летальный исход при встрече с коронавиурсом (т.к у них же будут антитела после вакцинации, и комплекс "вирус-антитело", садясь на этот рецептор, будет проникать внутрь макрофага и заражать его).
Это, конечно, спекуляции- но авторы обнаружили, что легочные макрофаги  и родственные им миелоидные клетки крови были определенно инфицированы коронавирусом-   так что попытались это как-то пояснить.
Важно, что примерно треть объема вируса определялась в не-эпителиальных клетках легких.
У некоторых умерших была тромбоэмболия легких- и артерий, и диссименирущая.
Из-за проблем с легкими и воспаления, у всех были ишемические поражения сердца, вплоть до некротических, и гипертрофии желудочков (особенно часто- правого).
У половины исследованных была отечность мозга.
Группы клеток, содержащих коронавирусы, или единичные клетки, были обнаружены почти во всех органах и тканах, что исследовали (жкт, мозг, сердце, селезенка, почки, щитовидка, этс)- но редко, и вирус в них еще не особо размножался. Правда, все больные у них умерли примерно через 2 недели, так что вирус просто "не успел".
Процитирую "обсуждение результатов", тк оно может расширять представление о происходящем. Но и сами результаты тоже познавательны.

Adequate therapy of a virus infection initiated sickness require the proper understanding of not only the virus infection induced cell damage but also the effect of different host responses. As most victims of Covid-19 are in advanced age with fragile health and numerous co-morbidities, the delineation of the virus induced pathologies require patient cohorts where the major tissue damage and eventually the direct cause of death can be directly linked to the presence of the replicating virus. Sars-Cov2 is considered a respiratory virus with lung damage as leading pathology. Our cohort comprises individuals who succumbed to the devastating effects of virus initiated lung damage, the Covid-19 associated acute respiratory distress syndrome. The gross changes of the lung parenchyma was similar to cases reported by others (Suess & Hausmann, 2020)(Bradley et al., 2020)(Zhou et al., 2020)(Barton et al., 2020)(Martines et al., 2020)(Adachi et al., 2020). In this cohort however we found that most macroscopic and microscopic pathologies were observed in the lung parenchyma as opposed to the upper or lower airways. Our findings suggest that in these patients the main site of the virus replication is in the pulmonary alveoli and not the laryngeal, pharyngeal, tracheal or bronchus epithelia. The leading pathology that eventually destroys the lung respiratory functions appears to be the massive consolidation of the lung parenchyma. The process is initiated by virus replication in the pneumocytes leading to the desquamation of the alveolar epithelia and various levels of breach in the barriers between alveolar capillaries and the intra-alveolar space. With the increasing severity of this breach edema fluid, plasma and eventually whole blood is leaking out into the alveolar space. Initiation of the coagulation cascade leads to the accumulation of fibrin filaments and/or intra-alveolar coagulated blood. Importantly only a minority of peumocytes showed lytic virus replication but most pneumocytes in the affected area showed pronounced cytopathic effects in form of cytoplasmic swelling, vesicular degeneration and nuclear atypia. At the present it is unclear how much of the epithelial damage can be interpreted as a by-stander effect or as a sign of abortive infection with arrested virus replication. Systematic mapping of early viral proteins such as the intact or cleaved products of the ORF1a and 1b would be required to answer this question. It has been suggested that various cellular elements of the vasculature such as endothelia (Varga et al., 2020) or pericytes (He et al., 2020) could be direct targets of Sars-Cov2 infection or would be damaged by subsequent inflammatory cell infiltrates (Becker, 2020). We have never seen any sign of viral presence in any of these structures nor inflammatory reaction against vascular wall components in our cohort. On the other hand we clearly demonstrate various levels of cytopathic effects of endothelial cells in the vicinity of virus infected epithelial cells by immunohistochemistry and electron microscopy. We also show that local release of red blood cells into the alveolar space is linked to the presence of virus infected epithelial cells. We interpret the endothelial damage as a by-stander effect possibly caused by soluble factors released from the infected pneumocytes. One candidate for such an effect might be the product of ORF3a, a viropore protein that has been shown to induce apoptosis in non-infected cells in SARS (C. M. Chan et al., 2009) and Covid-19(Ren et al., 2020). Extensive neutrophil infiltration and NET formation was hypothesized to significantly contribute to the development of ARDS (Barnes et al., 2020). In our series neutrophils were rarely present in the lethal ARDS lesions. All victims had high pre-mortem D-dimer levels in the serum similarly as reported in other studies (Mucha et al., 2020). High D-dimer together with well circumscribed radiological lesions regularly raises the clinical diagnosis of embolization of lung arteries. This diagnostic hypothesis was seemingly supported by pathology reports describing widespread intravascular fibrin micro thrombi implying generalized coagulopathy in Covid-19 patients (Calabrese et al., 2020)(Bradley et al., 2020)(Ackermann et al., 2020)(Vasquez-Bonilla et al., 2020)(Wichmann et al., 2020) . In our series we had only one case with disseminated intravascular coagulation. One other case with numerous pulmonary arterial thromb-embolization was shown to be composed mostly of platelet thrombi. Our data is congruent with the notion of the presence of extensive coagulation events. We suggest however that in many severe cases the bulk of the coagulum may be situated in the intra-alveolar rather than in the intravascular space. The increased frequency of CD61 positive intrapulmonary megakaryocytes along with the numerous thrombocyte micro-trombi raises the question if thrombocyte aggregation inhibitor therapy might be more beneficial for advanced Covid-19 patients than the routinely employed anticoagulation therapy. The reason of intra-alveolar bleeding appears to be linked to ill-defined cytopathic effects on vascular endothel in the vicinity of virus infected cells. It has been suggested that many of the pathological events of severe Covid-19 disease could be explained by virus induced cytokine storm characterized by increased levels of TNFalfa, IL-6, IL-8, IL-1beta and subsequent damage to the vascular endothelium (Huang et al., 2020). Recent data on severe Covid-19 ARDS cases showed much less pronounced cytokine elevations as compared to septic shock associated ARDS (Kox et al., 2020). Our data showing a great accumulation of CD163 positive myeloid cells in the lung consolidations and hilar lymph nodes in parallel with CD8 T, NK and B cell depletions in these tissues are in line with previous suggestions of M2 polarized macrophage induced effector T cell depletion (Liu et al., 2020)(Schulte-Schrepping et al., 2020). The elevated level of CD163 with the subsequent shedding is an emerging marker of virus infection associated macrophage activation syndrome with dismal outcome (McElroy et al., 2019) (Loomba et al., 2020).
The lung parenchymal consolidation was marked by a vigorous proliferation activity in the lung epitelium, endothelium, stroma and myelo-monocytic compartment. All these activities appear to compromise normal respiratory functions thus general proliferation inhibition with cytostatic chemicals or radiation might have a therapeutic value. Similarly, inhibition of the vigorous neoangiogenesis with anti-VEGF antibodies might be justified. The presence of the virus was reported in several tissues in various studies, mainly based on RT-PCR based detection (Hanley et al., 2020) or replication competence in ex vivo cultures (Hui et al., 2020). Already the eaƌliest studies desĐƌiďed of ͞ǀiƌal RNAŵLJ͟ iŶ the ďlood of soŵe of the ǀiƌus Đaƌƌieƌs (Huang et al., 2020). Our systematic mapping revealed a number of scattered cells in most organs without direct evidence of local virus replication outside of the alveolar space. Most cellular RNA degrades after 48 hours postmortem thus making virus detection in infected cells by RNA in situ hybridization very difficult. The virus however can be detected from infected lungs up to twelve days by RT-PCR (Edler et al., 2020) suggesting that the viral RNA persists well protected in the viral capsids. Our data show that extensive deposition of virus RNA in the hyaline membranes raises the possibility that released viral particles may be embedded in large quantities in the protective matrix of the hyaline membrane. Detection of hyaline membrane fragments in the upper airways may provide an alternative route for viral spreading. We suggest that the intra-alveolarly produced virus might leave the body with the exhaled air deposited in the proteinaceous matrix of the hyaline membrane. This matrix may provide extra environmental protection and possibly prolong the survival of the virus ex vivo. Both Sars-Cov (K. H. Chan et al., 2011) and Sars-Cov2 (Doremalen, Bushmaker, 2020) may remain viable on various surfaces for days when sprayed out as cell culture supernatant. Virus entrapped in dried out protein matrix may be prevented with receptor expressing target cells unless it is actively released. Inhaled virus bearing hyaline membrane particles are likely ingested by alveolar macrophages where the acidic environment and protease activity in the endosomes might initiate the generation of Spike fusion complex. Our data show that the frequent presence of viral RNA in CD68 positive macrophages is congruent with the hypothesis that phagocytic cells consuming virus carrying cellular debris might themselves become infected. This would provide an alveolus to alveolus transmission route through aerosol spreading that does not require replication events in ACE2 expressing cells. The absence of replicating virus in the upper airways and the focal, patchy, mosaic like involvement of the terminal lobules is also consistent with this type of viral spread. Alternative way for Sars-Cov2 entry into macrophages is through antiviral antibodies that may function as adapters through Fc receptor mediated viral binding and receptor mediated endocytosis. Macrophage infection through Fc receptors can lead to antibody dependent enhancement of the disease (ADE) that is well documented in vaccination models of dengue (Srikiatkhachorn & Yoon, 2016), feline infectious peritonitis (Takano et al., 2008)(Kipar & Meli, 2014) and experimental Sars vaccine models (Luo et al., 2018). The argument that Covid-19 vaccine would be free of ADE is largely based on the data that macrophages are refractory to Sars-Cov2 infection in vitro when cultured virus is used as infective agent. The demonstration of Spike RNA in CD68 positive cells in the infected lungs however raises the concern that certain vaccinated individuals who produce neutralizing antibodies against the Spike protein could succumb to ADE upon exposition to the circulating virus if the antibody would function as an adapter to bring the virus into the macrophages (Arvin et al., 2020)(Peeples, 2020)(Eroshenko et al., 2020). In vitro macrophage infection experiments using antibodies from victims who succumbed to Covid-19 will be required to test the validity of this scenario.
The added value of this study, beside detailed characterization of the lethal pulmonary injury caused by the virus, is the precise localization of the virus infected cells using RNA in situ hybridization in relation specific cell types identified by to immunohistochemical markers. Our data indicate massive bystander effect beyond the direct virus induced cytopathic damage. We show endothelium damage in the vicinity of virus replication without the presence of the virus in the endothelium. We present evidence of macrophage infection in the lungs. This finding can have major effect on the evaluation of the risk of ADE. We localize most of the virus production in the alveolar space and not in the upper airways, raising the possibility for alveolus to alveolus spread of the virus possibly embedded in hyaline membrane material. We show that in most fatal cases the fibrin coagulation happens in the intra-alveolar space and not in the vessels. We also show that thromboembolic events might be caused by increased thrombocyte aggregation rather than by increased intravascular coagulation. We show that massive proliferation activity in the epithelial, stromal, vascular and myelo-monocytic compartment contributes to the development of lung consolidations. Our data suggest that thrombocyte aggregation inhibition, angiogenesis inhibition and general proliferation inhibition may have a roll in the treatment of advanced Covid-19 ARDS.
Tags: здоровье, иммунитет, коронавирус, статьи
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