Additionally, 20 patients with bacterial sepsis-ARDS were included and served to compare blog of sinaling pathways the cytokine levels between the nvA(H1N1)-ARDS group and the bacterial sepsis-ARDS group.The study protocol was approved by the Ethics Committee for Clinical Research of the University of Medicine and Pharmacy ‘Iuliu Hatieganu’ Cluj Napoca and the hospital authority. Informed consent was obtained from each patient or their legal representative.The inclusion criteria were age > 16 years, symptoms compatible with influenza and confirmed nvA(H1N1) virus, bacterial severe sepsis with ARDS, and informed consent. The exclusion criteria were age < 16 years, known infection by human immunodeficiency virus, patients with other respiratory viral infections, bacterial sepsis without ARDS-syndrome, and refusal to consent.

The control group included 15 healthy volunteers without chronic or acute disease.Data were recorded prospectively by investigators at each hospital. The following data were recorded: age, sex, pregnancy, underlying diseases (chronic obstructive pulmonary disease, asthma, diabetes, chronic heart failure, chronic renal failure, cirrhosis, immunosuppression), obesity defined as body mass index > 30, and the time in days from symptom onset to hospital admission. Hematological, biochemical and microbiological results were included in the database. The extension of lung infiltrates on chest X-ray scan was registered as the number of quadrants involved. The severity and prognosis of the illness was assessed in adults using the Acute Physiology and Chronic Health Evaluation II (APACHE II) score and the Sepsis-related Organ Failure Assessment (SOFA) score.

ARDS was defined using the 1994 American-European Consensus Conference definitions [10]. The pulmonary dysfunction score was based on the PaO2:FiO2 ratio, ranging from 0 to 3 where grade 0 represented a ratio less or equal to 250; grade 1, a ratio ranging from 250 to 175; grade 2, a ratio ranging from 100 to 175; and grade 3, a ratio less or equal to 100[11].A(H1N1) influenza virus presence was confirmed by testing nasopharyngeal swabs or bronchoalveolar lavage specimens with real-time PCR (commercial kits: Full Velocity SYBR Green QRT-PCR/SuperScript III Platinum One-Step Quantitative RT-PCR Taqman; Invitrogen Corporation, Carlsbad, California, USA) at The National Influenza Centre of Cantacuzino Institute, Bucharest, Romania.

Cytokine and chemokine quantificationIn patients with nvA(H1N1)-mild disease, the serum samples were taken on hospital admission. In patients with nvA(H1N1)-ARDS infection, the serum samples were taken on admission to the ICU and 3 days later to determine cytokine kinetics. The installation of ARDS, either viral or Entinostat bacterial, in the course of the disease determined the time of admission to the ICU. In patients with bacterial sepsis-related ARDS, the serum samples were taken on admission to the ICU.