Intensive care for seriously ill patients affected by novel coronavirus sars - CoV - 2: Experience of the Crema Hospital, Italy.

AIMS: In this work, the survival and mortality data of 54 consecutive patients admitted to the Intensive Care Unit (ICU) and suffering from severe respiratory insufficiency imputable to viral SARS - CoV - 2 infection were analyzed and shared, after a critical review of the evidence in order to optimize the most dedicated clinical and treatment strategy, for a future 'targeted' management in the care of the possible return flu outbreak. METHODS: At our Emergency Department of the Crema Hospital, from the beginning of the pandemic until the end of June 2020, 54 consecutive patients admitted to ICU suffering from severe acute respiratory infection (SARI) and severe respiratory distress (ARDS) attributable to viral SARS - CoV - 2 infection were recruited. The recruitment criterion was based on refractory hypoxia, general condition and clinical impairment, comorbidities and CT images. The incoming parameters of the blood chemistry and radiology investigations and the timing of the gold - tracheal intubation were compared. Medical therapy was based on the application of shared protocols. RESULTS: The onset of symptoms was varyng, i.e. within the range of 1-14 days. The average time from the admission to the emergency room to the admission to intensive care was approximately 120 h. The average number of days of hospitalization in the ICU was 28 days. With a majority of male patients, the most significant age group was between 60 and 69 years. There were 21 deaths and, compared to the survivors, the deceased ones were older at an average age of about 67 years (vs an average age of the survivors of about 59 years). From the available data entering the ICU, the surviving patients presented average better values of oximetry and blood gas analysis, with a lower average dosage of D-Dimer than the deceased. Ones with a presence of bilateral pneumonia in all patients, the worsening of the ARDS occurred in 31 patients. 9 out of 25 patients early intubated died, while 12 out of 23 patients died when intubation was performed after 24 h of non-invasive ventilation. The presence of multiple comorbidities was shown in 17 of 28 patients and revealed an additional adverse prognostic factor. Also, more than one complication in the same patient were detected; after respiratory worsening, renal failure was more frequently found in 16 patients. Some particular complications such as lesions induced by ventilation with barotrauma mechanism (VILI), ischemic heart disease and the appearance of central and peripheral neurological events were detected too. CONSIDERATIONS: SARS - CoV - 2 disease is caused by a new coronavirus that has its main route of transmission through respiratory droplets and close contact, resulting in a sudden onset of the clinical syndrome with acute respiratory infection (SARI) and severe respiratory distress (ARDS). But it can also appear with other symptoms such as gastrointestinal or neurological events, as to be considered as a disease with multisystem phenotype. This pathology evolves towards a serious form of systemic disease from an acute lung damage to venous and arterial thromboembolic complications and multi-organ failure, mostly associated with high mortality. All patients received empirical or targeted antibiotic therapy for prevention and control of infections of potential pathogens, together with low molecular weight heparin therapy. The majority of patients was subjected to the off - label protocol with antivirals and hydroxychloroquine therapy, we used cortisone support therapy under surveillance and in 3 cases the protocol with anti - IL6 monoclonal antibody (Tolicizumab). In a simplified classification of the tomographic examination of the chest, mostly 3D and 2C lesions were found in the deceased patients with a prevalence of severe and moderate forms, whilst in the survivors the distribution appears with a prevalence of medium and moderate forms. Among the intubated patients, 21 patients, all suffering from worsening ARDS, died whilst there was no mortality in patients subjected to non-invasive ventilation it so. The heterogeneity of the respiratory syndromes and the presence of multiple comorbidities represent an unfortunate prognostic factor. Among the complications, besides the respiratory worsening, renal failure, liver failure and the state of sepsis were most frequently found; less frequent complications were lesions induced by ventilation with a barotrauma mechanism, ischemic heart disease, the appearance of central neurological events of sensory alterations, meningo - encephalitis and cerebral hemorrhage, and peripheral neurological events with polyneuro - myopathies. Mechanical ventilation can adversely affect the prognosis due to lung damage induced, protective ventilation remains the necessary treatment during severe hypoxia in patients with SARS - CoV - 2. The essential prerequisite remains the search for optimal 'customized' values since conditions can vary from patient to patient and, in the same patient, during different times of ventilation. CONCLUSIONS: In these extraordinary circumstances, our reality was among the most affected and was able to hold the impact thanks to the immediate great response set in place by the operators, although it costed us an effort especially the one to try to guarantee a high quality level of assistance and care compared to the huge wave of patients in seriously bad conditions. Further research on this heterogeneous pathology and data sharing could help identify a more dedicated clinical decision-making and treatment pathway that, together with a resource planning, would allow us to better face any new disease outbreak.

Detection of HCV antigens in liver graft: relevance to the management of recurrent post-liver transplant hepatitis C.

The aim of this study was to evaluate how the immunohistochemical detection of liver hepatitis C virus (HCV) antigens (HCV-Ag) could support the histologic diagnosis and influence the clinical management of post-liver transplantation (LT) liver disease. A total of 215 liver specimens from 152 HCV-positive patients with post-LT liver disease were studied. Histologic coding was: hepatitis (126), rejection (34), undefined (24; coexisting rejection grade I and hepatitis), or other (31). The percentage of HCV-Ag infected hepatocytes were evaluated, on frozen sections, by an immunoperoxidase technique. HCV-Ag were detectable early in 57% of cases within 30 days post-LT, 92% of cases between 31 and 180 days, and 74% of cases after more than 180 days. Overall, HCV-Ag were detected more frequently in histologic hepatitis as compared to rejection (P < 0.0001) with a higher percentage of positive hepatocytes (P < 0.00001). In 16 patients with a high number of HCV-Ag-positive hepatocytes (65%; range 40-90%) a clinical diagnosis of recurrent hepatitis (RHC) was made despite inconclusive histopathologic diagnosis. Multivariate analysis identified the percentage of HCV-Ag-positive hepatocytes and the time post-LT as independent predictors for RHC (P = 0.008 and P = 0.041, respectively) and the number of HCV-Ag-positive hepatocytes >/=50% as the only independent predictor for nonresponse (P < 0.001) in 26 patients treated with alpha-interferon plus ribavirin. In conclusion, HCV reinfection occurs early post-LT, reaching its peak within 6 months. Immunohistochemical detection of post-LT HCV reinfection support the diagnosis of hepatitis when the histologic features are not conclusive. A high number of infected cells, independently from the genotype, represents a negative predictive factor of response to antiviral treatment.

Clinical impact of non-organ-specific autoantibodies on the response to combined antiviral treatment in patients with hepatitis C.

BACKGROUND: Hepatitis C virus (HCV)-related chronic hepatitis is frequently associated with non-organ-specific autoantibodies (NOSAs), but available data about the relationship between NOSA positivity and the effect of antiviral therapy in persons with hepatitis C are few and controversial. Our aim was to evaluate the impact of NOSA positivity on the outcome of combined antiviral therapy in HCV-positive patients. METHODS: A total of 143 consecutive adult patients with hepatitis C were studied. Antinuclear antibody (ANA), anti-smooth muscle antibody (SMA), and anti-liver/kidney microsomal antibody type 1 (LKM1) were detected by indirect immunofluorescence. All patients were treatment naive and received combined antiviral therapy (interferon [IFN]-ribavirin) after enrollment in the study. Patients were classified as nonresponders if HCV RNA was detectable after 6 months of therapy, as relapsers if abnormal transaminase levels and reactivation of HCV replication were observed after the end of treatment, and as long-term responders if transaminase levels were persistently normal and HCV RNA was undetectable 6 months after the end of treatment. RESULTS: Thirty-seven patients (25%) were NOSA positive (SMA was detected in 19 patients, ANA in 10, ANA and SMA in 4, LKM1 in 3, and SMA and LKM1 in 1). The prevalence of long-term response was similar between NOSA-positive patients and NOSA-negative patients (48.6% vs. 56.6%; P=not significant). Compared with HCV genotype 1 (HCV-1), HCV genotypes other than 1 were more often associated with long-term response among NOSA-positive patients (93.3% vs. 30%; P=.0017). The overall rate of long-term response, irrespective of NOSA status, was 54.5%. Detection of HCV-1 and elevated gamma-glutamyl transpeptidase serum levels were independent negative prognostic factors of treatment response (P=.007 and P=.026, respectively). CONCLUSIONS: Combined antiviral treatment (IFN-ribavirin) is safe and effective in NOSA-positive patients with hepatitis C, even if long-term response is less likely in those infected with HCV-1.

Detection of recipient's cells in liver graft using antibodies to mismatched HLA class I antigens.

Engraftment by recipient's (R) cells has been already demonstrated in gender mismatched liver grafts using fluorescence in situ hybridization (FISH), with contrasting results concerning epithelial cells. Mismatch for human leukocyte antigen (HLA) class I (HLA-I) is quite common in patients with orthotopic liver transplantation (OLT). We thus aimed to assess whether monoclonal antibodies (MoAbs), currently employed in the HLA typing process, could be used to study the dynamics of R cells in liver grafts. A total of 50 frozen liver biopsies from 37 patients receiving a HLA mismatch liver were tested. Biopsies were obtained from 3 days to more than 360 days after OLT. Frozen sections of graft biopsies were stained using an immunoperoxidase technique with the proper MoAbs. In selected cases, a double immunofluorescence was also performed. Circulating R blood cells and sinusoidal cells were occasionally observed in liver biopsies obtained within 10 days after OLT and were commonly detected after 1 month. The number of sinusoidal cells continued to increase up to 6 months, as shown on serial biopsies. On the whole, R blood cells and R sinusoidal cells were detected in 86% and 82% of the biopsies, respectively. R hepatocytes and biliary cells were detected after 40 and 60 days after OLT, respectively, in 14% (hepatocytes), 8% (bile ducts), and 12% (proliferating bile ducts) of the biopsies. R hepatocytes presented as single cells or groups of few cells; their number was lower than 1% and apparently did not increase with time after OLT. In conclusion, it is possible to detect R cells in liver graft using MoAbs to specific mismatched HLA-I alleles. R sinusoidal cells start to appear after 10 days and are commonly observed after 1 month; bile duct cells and hepatocytes appear later and their number does not increase with time. Engraftment by R epithelial cells seems to be less important than previously reported.

Detection of the M30 neoepitope as a new tool to quantify liver apoptosis: timing and patterns of positivity on frozen and paraffin-embedded sections.

One of the first stages of apoptosis is cytokeratin cleavage mediated by caspases, which is associated with the expression of a neoepitope, the cleavage site of cytokeratin 18, identifiable by the M30 monoclonal antibody. The aim of this study was to evaluate the timing of neoantigen expression and its modifications in the various morphologic stages of apoptosis on frozen and paraffin-embedded sections from liver biopsies of patients with chronic hepatitis or transplanted liver. The appearance of this neoepitope coincides with the gradual disappearance of cytokeratins, with the appearance of nuclear DNA fragmentation, and with the presence of Councilman bodies. The staining patterns on paraffin-embedded sections of liver specimens were similar to those found in frozen sections, with a reduced sensitivity. The M30 antibody is correlated with apoptosis, and its specificity for epithelial cells makes this method the first choice for routine evaluation of apoptosis in liver epithelial cells.

Timing of reinfection and mechanisms of hepatocellular damage in transplanted hepatitis C virus-reinfected liver.

Pathogenic mechanisms and dynamics of hepatitis C virus (HCV) reinfection in orthotopic liver transplantation (OLT) are poorly defined. This study focuses on these aspects by studying 55 frozen biopsy specimens from transplant recipients with various histological diagnoses obtained from 4 days to 4 years post-OLT and 10 patients with HCV-related chronic hepatitis. The percentage of HCV-infected hepatocytes, number and distribution of CD8 and natural killer cells, and rates of hepatocellular apoptosis and proliferation were quantified by immunohistochemistry. HCV antigens were detected in 37% of biopsy specimens obtained within 20 days and 90% of biopsy specimens obtained from 21 days to 6 months after OLT. The number of HCV-infected hepatocytes was never less than 40% in acute hepatitis specimens and never greater than 30% in the other cases. Hepatocellular apoptosis was high in biopsy specimens of acute hepatitis and moderate in those from transplant recipients with normal histological characteristics, but still greater than in specimens of chronic active hepatitis. Proliferation correlated significantly with apoptosis. Lymphocyte infiltration was high and similar among cases of acute hepatitis, chronic hepatitis, and rejection. These data: (1) show that the detection of liver HCV antigens is sensitive enough to be used in clinical practice as a diagnostic tool to detect infection of the transplanted liver and might be useful, combined with conventional histological evaluation to detect hepatitic damage, for therapeutic decision making; (2) suggest direct cytotoxicity of HCV, as well as immunologic mechanisms possibly prevalent in chronic hepatitis and rejection, at least in the phase of acute massive liver infection; and (3) show that hepatocellular apoptosis and regeneration might be active enough to lead to replacement of the entire transplanted liver in 2 weeks.