Diagnostic value of white blood cell parameters for COVID-19: Is there a role for HFLC and IG?

INTRODUCTION: As the Coronavirus disease 2019 (COVID-19) pandemic is still ongoing with patients overwhelming healthcare facilities, we aimed to investigate the ability of white blood cell count (WBC) and their subsets, high fluorescence lymphocyte cells (HFLC), immature granulocyte count (IG), and C-reactive protein (CRP) to aid diagnosis of COVID-19 during the triage process and as indicators of disease progression to serious and critical condition. METHODS: We collected clinical and laboratory data of patients, suspected COVID-19 cases, admitted at the emergency department of University General Hospital of Ioannina (Ioannina, Greece). We selected 197 negative and 368 positive cases, confirmed by polymerase chain reaction test for severe acute respiratory syndrome coronavirus 2. COVID-19 cases were classified into mild, serious, and critical disease. Receiver operating characteristic curve and binary logistic regression analysis were utilized for assessing the diagnosing ability of biomarkers. RESULTS: WBC, neutrophil count (NEUT), and HFLC can discriminate efficiently negative cases from mild and serious COVID-19, whereas eosinopenia and basopenia are early indicators of the disease. The combined WBC-HFLC marker is the best diagnostic marker for both mild (sensitivity: 90.6% and specificity: 64.1%) and serious (sensitivity: 90.3% and specificity: 73.4%) disease. CRP and Lymphocyte count are early indicators of progression to serious disease whereas WBC, NEUT, IG, and neutrophil-to-lymphocyte ratio are the best indicators of critical disease. CONCLUSION: Lymphopenia is not useful in screening patients with COVID-19. HFLC is a good diagnostic marker for mild and serious disease either as a single marker or combined with WBC whereas IG is a good indicator of progression to critical disease.

Synthesis and characterization of the diastereomers Lambda- and Delta-[Ru(bpy)2(m-bpy-L-Arg-Gly-L-Asn-L-Ala-L-His-L-Glu-L-Arg)]Cl2 1H NMR studies on their interactions with the deoxynucleotide duplex d[(5'-GCGCTTAAGCGC-3')2] and d[(5'-CGCGATCGCG-3')2].

The diastereomeric complexes Lambda- and Delta-[Ru(bpy)(2)(m-bpy-7p)]Cl(2), (bpy=2,2'-bipyridine, m-bpy-7p=4-methyl-4'-Arg-Gly-Asn-Ala-His-Glu-Arg-CONH(2)-2,2'-bipyridine) were synthesized and characterized and their binding properties to the deoxynucleotide duplexes d(5'-CGCGATCGCG-3')(2) and d(5'-GCGCTTAAGCGC-3')(2) were studied by means of (1)H NMR spectroscopy. 7p is part of the recognition loop of the restriction endonuclease MunI, a type II restriction enzyme from Mycoplasma unidentified which recognizes the palindromic hexanucleotide sequence C/AATTG and cleaves it as indicated by the slash. The Delta-isomer binds to the terminal CG/GC major groove of d(CGCGATCGCG)(2) decanucleotide, whereas the Lambda-isomer approaches the GCT/CGA sequence. On the other hand, weak binding of the Delta-isomer to the end of d(GCGCTTAAGCGC)(2) into two different orientations is observed. In the case of the Lambda-isomer, the bpy ligand(s) are located into the major groove of the central TT/AA sequence. The role of appended peptide sequences in sequence selectivity binding to DNA is being addressed.

Copper(II) and zinc(II) complexes of the peptides Ac-HisValHis-NH2 and Ac-HisValGlyAsp-NH2 related to the active site of the enzyme CuZnSOD.

Copper(II) and zinc(II) complexes of the peptides Ac-HisValHis-NH2 and Ac-HisValGlyAsp-NH2 related to the active site of the enzyme CuZnSOD were studied by potentiometric and spectroscopic (UV-Vis, CD and EPR) techniques. The results reveal that both ligands have effective metal binding sites, but the tripeptide is a much stronger complexing agent than the tetrapeptide. The formation of a macrochelate via the coordination of the imidazolyl residues is suggested in the copper(II)-Ac-HisValHis-NH2 system in the acidic pH range, while a 4N complex predominates at physiological pH. The interaction of Ac-HisValHis-NH2 with zinc(II) results in the formation of a precipitate indicating polynuclear complex formation. Both copper(II)-Ac-HisValHis-NH2 and copper(II)-HisValHis systems exhibit catalytic activity toward the dismutation of superoxide anion at physiological pH, but the saturated coordination sphere of the metal ions in both systems results in low reactivity as compared to the native enzyme.

Copper(I) interaction with model peptides of WD6 and TM6 domains of Wilson ATPase: regulatory and mechanistic implications.

With the aim to investigate the mechanism of Cu(I) transport by Wilson ATPase (ATP7B), we have studied the interaction of the peptides 2K10p (CH(3)CO-Lys-Gly-Met-Thr-Cys-Ala-Ser-Cys-Val-His-Asn-Lys-CONH(2)), and 2K8p (CH(3)CO-Lys-Leu-Cys-Ile-Ala-Cys-Pro-Cys-Ser-Lys-CONH(2)), part of the sixth metal binding domain (WD6) and the sixth transmembrane segment (TM6) of Wilson ATPase, respectively, by means of CD, NMR spectroscopy and homology modeling. In addition, the interaction of Cu(I) with the 2K8p mutants 1s (CH(3)CO-Lys-Leu-Ser-Ile-Ala-Cys-Pro-Cys-Ser-Lys-CONH(2)), 2s (CH(3)CO-Lys-Leu-Cys-Ile-Ala-Ser-Pro-Cys-Ser-Lys-CONH(2)) and 3s (CH(3)CO-Lys-Leu-Cys-Ile-Ala-Cys-Pro-Ser-Ser-Lys-CONH(2)), containing two cysteines in various positions, have been studied with the same methods, in order to understand the role of each cysteine in copper binding. Our studies show that the three cysteine thiolates present in the 2K8p peptide sequence act mainly as bridging ligands for Cu(I) binding, and dithiothreitol acts as an important ligand in Cu(I) ligation by 2K10p and the 2K8p mutants. Formation of oligomeric species has been evidenced for all peptides except 2s. Shift of the equilibrium between the various oligomeric species has been accomplished by reducing the Cu(I):peptide ratio. Significant shifts of proline protons upon interaction with Cu(I) have been observed for all proline containing peptides implying a possible role of proline in facilitating Cu(I) binding. These findings have been further discussed with respect to the molecular basis of copper trafficking and intermolecular interactions.

NMR analysis of duplex d(CGCGATCGCG)2 modified by Lambda- and Delta-[Ru(bpy)2(m-GHK)]Cl2 and DNA photocleavage study.

The interaction of the diastereomeric complexes Lambda-[Ru(bpy)2(m-GHK)]Cl2 and Delta-[Ru(bpy)2(m-GHK)]Cl2 (bpy is 2,2'-bipyridine, GHK is glycine-L-histidine-L-lysine) with the deoxynucleotide duplex d(5'-CGCGATCGCG)2 was studied by means of 1H NMR spectroscopy. At a Delta-isomer to DNA ratio of 1:1, significant shifts for the metal complex are observed, whereas there is negligible effect on the oligonucleotide protons and only one intermolecular nuclear Overhauser effect (NOE) is present at the 2D nuclear Overhauser enhancement spectroscopy spectrum. The 1Eta NMR spectrum at ratio 2:1 is characterized by a slight shift for the Delta-isomer's bpy aromatic protons as well as significant shifts for the decanucleotide G4 H1' and Eta2'', A5 H2, G10 H1', T6 NH and G2 NH protons. Furthermore, at ratio 2:1, 11 intermolecular NOEs are observed. The majority of the NOEs involve the sugar Eta2' and Eta2'' protons sited in the major groove of the decanucleotide. Increasing the Delta-isomer to d(CGCGATCGCG)2 ratio to 5:1 results in noteworthy spectral changes. The Delta-isomer's proton shifts are reduced, whereas significant shifts are observed for the decanucleotide protons, especially the sugar protons, as well as for the exchangeable protons. Interaction is characterized by the presence of only one intermolecular NOE. Furthermore, there is significant broadening of the imino proton signals as the ratio of the Delta-isomer to DNuAlpha increases, which is attributed to the opening of the two strands of the duplex. The Lambda-isomer, on the other hand, approaches the minor groove of the oligonucleotide and interacts only weakly, possibly by electrostatic interactions. Photocleavage studies were also conducted with the plasmid pUC19 and a 158-bp restriction fragment, showing that both diastereomers cleave DNA with similar efficiency, attacking mainly the guanines of the sequence probably by generating active oxygen species.

Interaction of Cu(II)with His-Val-Gly-Asp and of Zn(II) with His-Val-His, two peptides at the active site of Cu,Zn-superoxide dismutase.

His-Val-His and His-Val-Gly-Asp are two naturally occurring peptide sequences, present at the active site of Cu,Zn-superoxide dismutase (Cu,Zn-SOD). We have already studied the interaction of His-Val-His=A (copper binding site) with Cu(II) and of His-Val-Gly-Asp=B (zinc binding site) with Zn(II). As a continuation of this work and for comparison purposes we have also studied the interaction of Zn(II) with His-Val-His and Cu(II) with His-Val-Gly-Asp using both potentiometric and spectroscopic methods (visible, EPR, NMR). The stoichiometry, stability constants and solution structure of the complexes formed have been determined. Histamine type of coordination is observed for/ZnAH/(2+), /ZnA/(+), /ZnA(2)H/(+) and/ZnA(2)/ in acidic pH while deprotonation of coordinated water molecules is observed at higher pH. /CUB/ species is characterized by the formation of a macrochelate and histamine type coordination. Its stability results in the suppression of amide deprotonation which occurs at high pH resulting in the formation of the highly distorted from square planar geometry 4N complex/CuBH(-3)/(3).