Predictive values of immune indicators on respiratory failure in the early phase of COVID-19 due to Delta and precedent variants.

Background: Immune response indicators in the early phase of COVID-19, including interferon and neutralizing responses against SARS-CoV-2, which predict hypoxemia remains unclear. Methods: This prospective observational study recruited patients hospitalized with COVID-19 (before emergence of omicron variant). As the immune indicators, we assessed the serum levels of IFN-I/III, IL-6, CXCL10 and VEGF, using an ELISA at within 5 days after the onset of symptoms, and serum neutralizing responses using a pseudovirus assay. We also assessed SARS-CoV-2 viral load by qPCR using nasal-swab specimens and serum, to assess the association of indicators and viral distribution. Results: The study enrolled 117 patients with COVID-19, of which 28 patients developed hypoxemia. None received vaccine before admission. Serum IFN-I levels (IFN-α and IFN-ß), IL-6, CXCL10, LDH and CRP were significantly higher in patients who developed hypoxemia. A significant association with nasopharyngeal viral load was observed only for IFN-I. The serum levels of IFN-α, IL-6, CXCL10 were significantly associated with the presence of RNAemia. Multivariable analysis showed higher odds ratio of IFN-α, with cut-off value of 107 pg/ml, in regard to hypoxemia (Odds ratio [OR]=17.5; 95% confidence interval [CI], 4.7-85; p<0.001), compared to those of IL-6, >17.9 pg/ml (OR=10.5; 95% CI, 2.9-46; p<0.001). Conclusions: This study demonstrated that serum IFN-α levels in the early phase of SARS-CoV-2 infection strongly predict hypoxemic respiratory failure in a manner different from that of the other indicators including IL-6 or humoral immune response, and instead sensitively reflect innate immune response against SARS-CoV-2 invasion.

Hyperactivation of the RAS signaling pathway in myelodysplastic syndrome with AML1/RUNX1 point mutations.

AML1/RUNX1 mutations have been reported frequently in myelodysplastic syndrome (MDS) patients, especially those diagnosed with refractory anemia with excess blast (RAEB), RAEB in transformation (RAEBt), or AML following MDS (these categories are defined as MDS/AML). Although AML1 mutations are suspected to play a pivotal role in the development of MDS/AML, acquisition of additional genetic alterations is also necessary. We analyzed gene alterations in MDS/AML patients with AML1 mutations, comparing them to alterations in those without an AML1 mutation. AML1 mutations were significantly associated with -7/7q-, whereas MDS/AML patients without AML1 mutations showed a high frequency of -5/5q- and a complex karyotype. Patients with AML1 mutations showed more mutations of their FLT3, N-RAS, PTPN11, and NF1 genes, resulting in a significantly higher mutation frequency for receptor tyrosine kinase (RTK)-RAS signaling pathways in AML1-mutated MDS/AML patients compared to AML1-wild-type MDS/AML patients (38% versus 6.3%, P < 0.0001). Conversely, p53 mutations were detected only in patients without AML1 mutations. Furthermore, blast cells of the AML1-mutated patients expressing surface c-KIT, and SHP-2 mutants contributed to prolonged and enhanced extracellular signal-regulated kinase activation following stem cell factor stimulation. Our results suggest that MDS/AML arising from AML1/RUNX1 mutations has a significant association with -7/7q- alteration, and frequently involves RTK-RAS signaling pathway activation.

Antiangiogenic activity of curcumin in hepatocellular carcinoma cells implanted nude mice.

Antiangiogenic activity of curcumin on the tumor neogenesis was investigated by evaluating the density of neocapillaries induced by Hepatocellular carcinoma cells (HepG2) in mice, using intravital fluorescence videomicroscopy. Male BALB/c nude mice (20-25 g) were used, and a dorsal skin-fold chamber was implanted. HepG2 (30 microl of 2 x 10(6) cells) were inoculated on the upper surface of the skin within the chamber. The mice were divided into two groups as follows. Dimethyl sulfoxide solution (0.1%) was fed (HepG2 group, n=5) or curcumin solution (3000 mg/kg bw) was fed oral daily (HepG2-Cur group, n=5), one day after the inoculation of HepG. On days 7 and 14 post-tumor-inoculation, the tumor microvasculature was visualized by injecting 0.1 ml of 0.5% rhodamine B isothiocyanate-labeled dextran intravenously, and observed under an intravital fluorescence videomicroscope. Based on the recorded videoimage, the tumor neocapillary density and microvasculature were evaluated using a digital image analysis and correlated with the tumor area. The image analysis demonstrated that in the HepG2-group the neocapillary densities were significantly increased on day 7, and day 14, compared to the aged-matched Sham-group (P<0.05). In the HepG2-Cur group, the increase of tumor neocapillary density was attenuated significantly. It was suggested that high dose of curcumin might be an effective anti-angiogenic drug in the treatment against tumor.

Quantitative assessment of cerebral neocapillary network and its remodeling in mice using intravital fluorescence videomicroscopy.

To assess the responses of different growth factors on cerebral neocapillary density (NCD), cerebral angiogenesis was induced in mice using growth factors, basic fibroblast growth factor (bFGF) and platelet-derived growth factor (PDGF) at a concentration of 6 ng/ml each. Intravital fluorescence videomicroscopy was used to quantitatively evaluate microhemodynamic parameters such as diameter and red cell velocity. The gel-nylon mesh-sandwich system was implanted over the exposed cortex. After incubation for different periods of time (days 7, 14 or 28), fluorescein isothiocyanate (FITC)-labeled red cells were injected through a carotid artery and the neocapillaries on the upper surface of the nylon mesh were observed under a fluorescence videomicroscope. Based on the recorded videoimages, we evaluated the density, diameter and red cell velocity of the neocapillaries. The NCD in the bFGF group on day 7 was significantly higher than that in the PDGF group on day 7 (P < 0.01). The NCD (index) reached 100% on day 14, while it reduced significantly in both the groups on day 28. The neocapillary diameter was greater than that of the pre-existing capillaries on day 7. On day 14, a clear difference appeared in the capillary density between large and small vessels. The red cell velocity increased with the number of days after incubation. The response of cerebral neocapillaries to acetylcholine was measured after 28 days of incubation with growth factor bFGF and with PDGF. The red cell velocity increased significantly from its basal value in the PDGF group. These results suggest that the neocapillaries in the PDGF group matured earlier than those in the bFGF group.

Bronchiolitis obliterans with organizing pneumonia versus chronic eosinophilic pneumonia: high-resolution CT findings in 81 patients.

OBJECTIVE: The objective of this research was to compare high-resolution CT findings of bronchiolitis obliterans with organizing pneumonia (BOOP) with those of chronic eosinophilic pneumonia (CEP) and to determine whether high-resolution CT can differentiate the two. MATERIALS AND METHODS: We retrospectively reviewed high-resolution CT scans of 38 patients with BOOP and 43 patients with CEP. Without knowledge of the diagnosis, two radiologists evaluated the frequency and distribution of high-resolution CT findings in both groups of patients and made a diagnosis using a three-point scale of confidence. RESULTS: Nodules, nonseptal linear or reticular opacities, and bronchial dilatation were significantly more common in BOOP than in CEP (31.6% vs. 4.7%, p < 0.005; 44.7% vs. 9.3%, p < 0.001; and 57.9% vs. 25.6%, p < 0.005, respectively). Septal line thickening was more frequent in CEP than in BOOP (72.1% vs. 39.5%, p < 0.005). Peribronchial distribution of consolidation was more frequent in BOOP than in CEP (28.9% vs. 9.3%, p < 0.05). A correct diagnosis was made in 69.7% of cases, and the diagnostician was confident in 21.7%. Interobserver agreement was good (kappa = 0.6). CONCLUSION: Although several of the high-resolution CT findings of BOOP and CEP are different, these diseases are differentiated with confidence in only a small percentage of cases.

Cytotoxic T lymphocyte antigen 4 gene polymorphism confers susceptibility to type 1 diabetes in Japanese children: analysis of association with HLA genotypes and autoantibodies.

OBJECTIVE: Although the polymorphisms of the cytotoxic T lymphocyte antigen 4 (CTLA4) gene have been shown to be associated with Type 1 diabetes in Caucasians, some conflicting results have been reported among subjects of different ethnic backgrounds. We examined a CTLA4 polymorphism and its relationship to human leucocyte antigen (HLA) genotypes and autoantibodies for glutamic acid decarboxylase 65 (GAD65) and IA-2 in Japanese children with Type 1 diabetes. SUBJECTS AND MEASUREMENTS: The study group consisted of 125 childhood-onset Japanese subjects (50 males, 75 females) with Type 1 diabetes. The CTLA4 A/G polymorphism at position 49 was analysed using a PCR-restriction fragment length polymorphism (PCR-RFLP) method. The HLA-DRB1 and DQB1 genotypes were defined by DNA analysis using PCR-sequence-specific oligonucleotide (PCR-SSO) probes. The GAD65 autoantibody (GAD65Ab) and IA-2 autoantibody (IA-2Ab) titres were measured using radioimmunoassay. RESULTS: The distribution of genotype frequencies differs between subjects with Type 1 diabetes (GG: 46%, AG: 50%, AA: 5%) and controls (GG: 39%, AG: 44%, AA: 17%) (P < 0.01). The frequency of the G allele is higher in the diabetes group than in the controls (P < 0.05). When the subjects were subdivided according to HLA genotype, the two major HLA high-risk groups, with DR9-DQ9 and DR4-DQ4, that are unique to Japanese populations showed no difference in their CTLA4 polymorphism frequencies. Although no association between the CTLA4 polymorphism and the prevalence of GAD65Ab was found, CTLA4 GG subjects that had been newly diagnosed (< 9 months) had significantly higher levels of autoantibodies than AG subjects (P < 0.01). The prevalence and titres of IA-2Ab were not associated with the CTLA4 polymorphism. CONCLUSIONS: The CTLA4 gene might confer a susceptibility to childhood-onset Type 1 diabetes in the Japanese population. The association between this CTLA4 polymorphism and the HLA genotype was similar for both major groups with HLA high-risk alleles. CTLA4 might contribute to the humoral immune response to GAD in newly diagnosed subjects.

Expression of tumor necrosis factor-alpha protein in the myocardium in fatal myocarditis.

BACKGROUND: Tumor necrosis factor (TNF)-alpha is the most studied cytokine in the failing human heart and in experimental murine myocarditis. We have investigated the expression of TNF-alpha in the myocardium in human myocarditis. METHODS: We examined endomyocardial biopsy (n = 4) and autopsy (n = 5) tissues obtained from nine patients diagnosed with myocarditis by the Dallas criteria. Expression of TNF-alpha in the hearts was immunohistochemically studied using monoclonal antibodies against human TNF-alpha. RESULTS: Tumor necrosis factor-alpha protein was expressed in the myocardium of six of the nine patients studied. Four of five fatal patients showed intense immunoreactivity for TNF-alpha compared with survivors. Furthermore, left ventricular systolic function was reduced in patients with TNF-alpha-positive hearts. CONCLUSIONS: These findings may support the suggestion that TNF-alpha plays an important role in cardiac dysfunction and myocytic damage in fatal human myocarditis.

VEGF is causative for pulmonary hypertension in a patient with Crow-Fukase (POEMS) syndrome.

We report a case of Crow-Fukase (POEMS) syndrome associated with pulmonary hypertension (PH). In this case, the concentration of vascular endothelial growth factor (VEGF) was extremely high in the serum, and the levels of IL-1beta, IL-6, TNF-alpha, and thiamine, which were thought in past reports to be mediators of PH in Crow-Fukase syndrome, were normal. After prednisolone therapy, PH disappeared with a dramatic decrease in serum VEGF. Our results suggest that VEGF is closely correlated with PH in Crow-Fukase syndrome.

Asian traditional medicine: from molecular biology to organ circulation.

Asian tradition medicine (ATM), such as herbal medicine and acupuncture, has been widely used in the treatment of diseases, but the effects still remain unclear at the level of the microcirculation. Up to now, a variety of approaches have been made to comprehensively evaluate the significance of ATM. Microcirculatory studies of ATM are introduced with reference to the history of international workshops in Asia.

Microvascular vasodilatory responses to electric acupuncture in rat brain under acute hemorrhagic hypotension.

The present experiment attempted to evaluate the effect of electrical acupuncture on the cerebral microcirculation in anesthetized rats, using fluorescence videomicroscopy. Changes in the pial arteriolar diameter under acute hemorrhagic hypotension were examined quantitatively. The present results suggest that acupuncture may be effective in improving the cerebral microcirculation in hemorrhagic hypotension.

Effects of adrenomedullin on the cardiac performance and coronary flow in an isolated perfused rat heart model.

The effect of adrenomedullin (AM) on the cardiac performance and coronary flow were studied in an isolated perfused rat heart model based on the modified Langendorff method. The heart rate (HR), electrocardiogram (ECG), left ventricular contraction (LVC) (dP/dt), and coronary flow (CF) were measured before and after the application of AM. The effect of AM on the coronary flow was examined in the model with and without endothelial degradation, using different inhibitors such as N(G)-nitro-L-arginine, glibenclamide, and indomethacin. The present results indicated that AM increased HR and CF, but decreased LVC significantly, while it had no effect on ECG. The vasodilatory effect of AM was discussed in views of endothelial-dependence due to nitric oxide and K+ channel activation.

Microcirculatory characterization of cerebral angiogenesis in mice using intravital videomicroscopy.

The present study investigated microcirculatory characteristics of the cerebral neovasculature induced in mice, using basic fibroblast growth factor (bFGF) and platelet derived growth factor (PDGF). The nylon-mesh sandwich (collagen gel/growth factor in bovine serum albumin embedded in between two nylon-mesh pieces) was used to induce angiogenesis. After different days of incubation, the observations of neocapillaries were done on the upper surface of the nylon-mesh, using fluorescence video-microscopy. The neocapillary diameter, red cell velocity, and the neocapillary density were evaluated based on the video-image. The neocapillaries were visible on the upper surface of the mesh on the 6th day after the incubation, and red cells started to flow from the day 7. The neocapillary red cell velocity increased with days after incubation, but its level was less than that of the pre-existing capillaries. The neocapillary diameter decreased as the neocapillaries got matured. The neocapillary density was dependent on the doses of bFGF and PDGF. The neocapillary diameter did not alter with the higher concentration as well as with different growth factors. Both bFGF and PDGF showed an increase in red cell velocity at high concentration.

Microcirculatory responses to repeated embolism-reperfusion in cerebral microvessels of cat: a fluorescence videomicroscopic study.

Intra-carotid injections of degradable starch microspheres (DSMs) can induce DSM embolism-reperfusion in the level of cerebral arterioles. Vascular responses of cerebral arterioles to repeated DSM embolism (ischemia)-reperfusion were examined when DSMs were injected twice through a carotid artery with a time interval of 30 min. Arteriolar diameter was measured from images of arterioles filled with rhodamine-B isothiocyanate dextran and red cell velocity was measured with a dual window technique using FITC-labeled red cells as a flow tracer. DSM embolization induced ischemia (flow reduction including stasis) for approximately 10 min in the level of microvessels (arterioles). Cerebral arterioles began to dilate immediately after embolism induced by the DSM injection and vasodilation was sustained until reperfusion. After reperfusion the arteriole began to constrict and the arteriolar diameter returned to the initial diameter level at approximately 20 min after the DSM injection. The arteriolar diameters for the second DSM embolism showed a similar response to those for the first embolism in 7 out of 8 cats. It can be concluded that the vascular response of cerebral arterioles to the second embolism-reperfusion could not be affected by the first embolism-reperfusion.

The clinical usefulness of liquid human growth hormone (hGH) (Norditropin SimpleXx in the treatment of GH deficiency.

Human growth hormone (hGH) is an essential therapeutic drug for the treatment of GH deficiency. The development of recombinant GH using a pen injection system has enabled easy and safe treatment of GH-deficient patients; however, the process of dissolving hGH in the powder form is complicated and dangerous. In this study, we investigated the usefulness of a newly developed liquid form of hGH (Norditropin((R)) SimpleXx(TM)) in the treatment of 51 patients with GH deficiency. Fifteen previously untreated patients with GH deficiency were treated with liquid hGH (group A), and 36 patients who had previously used hGH in the powder form were changed to the liquid form (group B). Both groups were treated with liquid hGH 0.5 IU/kg per week for 6 months. The growth rate of patients in group A increased from 4.0 +/- 2.4 cm/year to 9.2 +/- 2.9 cm/year. The patients in group B continued to grow at the same rate as before using the liquid hGH therapy. Questionnaires to the patients in group B demonstrated that 85% preferred the convenience of using the new liquid form of hGH. Our results indicate that liquid hGH has similar efficacy to that of powder hGH, but its improved convenience may have a beneficial effect on patient compliance.

No enzyme activity of 25-hydroxyvitamin D3 1alpha-hydroxylase gene product in pseudovitamin D deficiency rickets, including that with mild clinical manifestation.

Pseudovitamin D deficiency rickets (PDDR) is an autosomal recessive disorder caused by defect in the activation of vitamin D. We recently isolated 25-hydroxyvitamin D3 1alpha-hydroxylase gene and identified four homozygous inactivating missense mutations in this gene by analysis of four typical cases of PDDR. This disease shows some phenotypic variation, and it has been suspected that patients with mild phenotypes have mutations that do not totally abolish the enzyme activity. To investigate the molecular defects associated with the phenotypic variation, we analyzed six additional unrelated PDDR patients: one with mild and five with typical clinical manifestation. By sequence analysis, all six patients were proven to have mutations in both alleles. The mutations varied, and we identified four novel missense mutations, a nonsense mutation, and a splicing mutation for the first time. The patient with mild clinical symptoms was compound heterozygous for T321R and a splicing mutation. The splice site mutation caused intron retention. Enzyme activity of the T321R mutant was analyzed by overexpressing the mutant 1alpha-hydroxylase in Escherichia coli cells to detect the subtle residual enzyme activity. No residual enzyme activity was detected in T321R mutant or in the other mutants. These results indicate that all of the patients, including those of mild phenotype, are caused by 1alpha-hydroxylase gene mutations that totally abolish the enzyme activity.