A turn-on fluorescent nanosensor for H2S detection and imaging in inflammatory cells and mice.

Hydrogen sulfide (H2S) is an endogenously generated gaseous signaling molecule and is known to be involved in the occurrence and development of inflammation. To better understand its physiological and pathological process of inflammation, reliable tools for H2S detection in living inflammatory models are desired. Although a number of fluorescent sensors have been reported for H2S detection and imaging, water-soluble and biocompatibility nanosensors are more useful for imaging in vivo. Herein, we developed a novel biological imaging nanosensor, XNP1, for inflammation-targeted imaging of H2S. XNP1 was obtained by self-assembly of amphiphilic XNP1, which was constructed by the condensation reaction of the hydrophobic, H2S response and deep red-emitting fluorophore with hydrophilic biopolymer glycol chitosan (GC). Without H2S, XNP1 showed very low background fluorescence, while a significant enhancement in the fluorescence intensity of XNP1 was observed in the presence of H2S, resulting in a high sensitivity toward H2S in aqueous solution with a practical detection limit as low as 32.3 nM, which could be meet the detection of H2S in vivo. XNP1 also has a good linear response concentration range (0-1 µM) toward H2S with high selectivity over other competing species. These characteristics facilitate direct H2S detection of the complex living inflammatory cells and drug-induced inflammatory mice, demonstrating its practical application in biosystems.

Construction of a robust turn-on fluorescence NIR sensor for rapid detection and imaging of ONOO- in inflammatory models.

Peroxynitrite (OONO-) is closely related to the occurrence and development of health and inflammatory diseases. The physiological and pathological results of OONO- are related to the local concentration of ONOO-. Therefore, to develop of a simple, rapid and reliable OONO- detection tool is badly needed. In this work, we developed a small-molecule near-infrared (NIR) turn-on fluorescence sensor (NN1), harnessing a well-known response group phenylboronic acid response toward OONO-. It shows high detection sensitivity and yields a ratio (I658/I0) fluorescence enhancement (∼280-fold). In addition, NN1 can be effectively used to detect endogenous and exogenous ONOO- in living inflammatory cells. Notably, NN1 can be applied to OONO- imaging analysis in drug-induced inflammatory mice model with satisfactory results. Therefore, NN1 is a robust molecular biological tool, which has a good prospect in the study of ONOO- and the occurrence and development of inflammatory diseases.

Diagnosis of prenatal 22q11.2 duplication syndrome: a two-case study.

The objective of the study was to perform the prenatal diagnosis of two foetuses with 22q11.2 duplication for 2.5 Mb after noninvasive prenatal testing (NIPT), and to explore the prenatal diagnosis and genetic characteristics of these foetuses. After amniocentesis, each foetus was diagnosed through karyotype analysis and single-nucleotide polymorphism array (SNP-array), and copy number variation using shotgun sequencing (CNV-seq) was carried out on each mother's peripheral blood for comparative analysis. Both pregnant woman 1 and pregnant woman 2 had foetal amniotic fluid chromosomal karyotypes of 46, XN. The SNP-array result for foetus 1 was arr[hg19] 22q11.21(18,648,856-21,800,471) x3; namely, 22q11.2 had a 3.1 Mb repeat, and the SNP-array result of foetus 2 was arr[hg19]22q11.2(18,648,855-21,464,764) x3; there was a 2.4 Mb repeat of 22q11.2. The CNV-Seq result of the peripheral blood of pregnant woman 1 was seq[hg19]22q11.2(18,953,139-21,449,967) x3; namely, in this mother's 22q11.2 region, there was ~2.5 Mb of duplicate fragment that was pathogenic to CNV. We confirmed that case 1 was inherited from the mother by CNV-seq. In both cases, however, there were key region deletions, including 41 OMIM genes such as CLTCL1, HIRA and TBX1. Both SNP-array and CNV-seq can effectively diagnose 22q11.2 duplication syndrome and clarify its fracture site and involved genes, which may facilitate understanding of the genotype and phenotype correlations.

Markers Associated With Tumor Recurrence in Patients With Breast Cancer Achieving a Pathologic Complete Response After Neoadjuvant Chemotherapy.

Background: Patients who achieve a tumor pathologic complete response (pCR) after neoadjuvant chemotherapy (NAC) have better outcomes than patients with residual tumor. However, tumors still recur in the pCR patients. Therefore, we aim to explore factors associated with tumor recurrence in this patient population. Methods: A total of 1,913 patients diagnosed with breast cancer between 1995 and 2020 and received NAC were included in this analysis. Clinicopathological data of the patients were retrospectively collected. We used Cox regression analysis to assess the associations of clinicopathological factors with patients' outcome. Proteomic study of tumors was applied to identify differentially expressed proteins (DEPs) between tumors from the pCR patients with tumor recurrence and tumors from those without tumor recurrence. PPI network analysis of the corresponding genes of DEPs was used to identify the hub genes. The prognostic value of the corresponding genes of DEPs was evaluated using two online databases, Kaplan-Meier Plotter and bc-GenExMiner. The genes that were significantly associated with patients' survival in both databases, as well as being identified as hub genes, were considered as potential prognostic markers for pCR patients. Publicly available data from Gene Expression Omnibus (GEO) was used to verify the prognostic value of the identified marker. Results: Among the 1,913 included patients, 420 had tumor pCR. The median follow-up for the pCR patients was 32.6 months (IQR, 16.3-55.5). Overall estimated 5-year risk of tumor recurrence for the pCR patients was 11%. Multivariable analysis showed that a higher pre-NAC clinical T stage and N stage were independent predictors for increased risk of tumor recurrence (hazard ratio [HR] 2.57, 95% confidence interval [CI] 1.01-6.51, P=0.047 for clinical T stage and HR 3.48, 95%CI 1.37-8.83, P=0.009 for clinical N stage). NAC regimens, the type of breast and axillary surgery, and adjuvant chemotherapy were not associated with tumor recurrence. Finally, aldehyde dehydrogenase (ALDH) 3A2 was identified by the proteomic study and was verified as a potential predictor for tumor recurrence in the pCR patients (with a median follow up of 3.78 years for dataset GSE32603 and 2.74 years for dataset GSE25066 from GEO, tumor recurrence rate: low versus high expression, 20.7% versus 4.5% [data from GSE32603]; 10.9% versus 0% [data from GSE25066]). Conclusions: Clinical T stage, clinical N stage and tumor expression of ALDH3A2 were potential markers for predicting tumor recurrence in the pCR patients after NAC.

Clinical and antibody characteristics reveal diverse signatures of severe and non-severe SARS-CoV-2 patients.

BACKGROUND: COVID-19 pandemic continues, clarifying signatures in clinical characters and antibody responses between severe and non-severe COVID-19 cases would benefit the prognosis and treatment. METHODS: In this study, 119 serum samples from 37 severe or non-severe COVID-19 patients from the First People's Hospital of Yueyang were collected between January 25 and February 18 2020. The clinical features, antibody responses targeting SARS-CoV-2 spike protein (S) and its different domains, SARS-CoV-2-specific Ig isotypes, IgG subclasses, ACE2 competitive antibodies, binding titers with FcγIIa and FcγIIb receptors, and 14 cytokines were comprehensively investigated. The differences between severe and non-severe groups were analyzed using Mann-Whitney U test or Fisher's exact test. RESULTS: Severe group including 9 patients represented lower lymphocyte count, higher neutrophil count, higher level of LDH, total bile acid (TBA) (P < 1 × 10-4), r-glutaminase (P = 0.011), adenosine deaminase (P < 1 × 10-4), procalcitonin (P = 0.004), C-reactive protein (P < 1 × 10-4) and D-dimer (P = 0.049) compared to non-severe group (28 patients). Significantly, higher-level Igs targeting S, different S domains (RBD, RBM, NTD, and CTD), FcγRIIa and FcγRIIb binding capability were observed in a severe group than that of a non-severe group, of which IgG1 and IgG3 were the main IgG subclasses. RBD-IgG were strongly correlated with S-IgG both in severe and non-severe group. Additionally, CTD-IgG was strongly correlated with S-IgG in a non-severe group. Positive RBD-ACE2 binding inhibition was strongly associated with high titers of antibody (S-IgG1, S-IgG3, NTD-IgG, RBD-IgA, NTD-IgA, and CTD-IgA) especially RBD-IgG and CTD-IgG in the severe group, while in the non-severe group, S-IgG3, RBD-IgG, NTD-IgG, and NTD-IgM were correlated with ACE2 blocking rate. S-IgG1, NTD-IgM and S-IgM were negatively associated with illness day in a severe group, while S-IgG3, RBD-IgA, CTD-IgA in the severe group (r = 0.363, P = 0.011) and S-IgG1, NTD-IgA, CTD-IgA in the non-severe group were positively associated with illness day. Moreover, GRO-α, IL-6, IL-8, IP-10, MCP-1, MCP-3, MIG, and BAFF were also significantly elevated in the severe group. CONCLUSION: Antibody detection provides important clinical information in the COVID-19 process. The different signatures in Ig isotypes, IgG subclasses, antibody specificity between the COVID-19 severe and non-severe group will contribute to future therapeutic and preventive measures development.

Systemic perturbations of key metabolites in diabetic rats during the evolution of diabetes studied by urine metabonomics.

BACKGROUND: Elucidation of metabolic profiles during diabetes progression helps understand the pathogenesis of diabetes mellitus. In this study, urine metabonomics was used to identify time-related metabolic changes that occur during the development of diabetes mellitus and characterize the biochemical process of diabetes on a systemic, metabolic level. METHODOLOGY/PRINCIPAL FINDINGS: Urine samples were collected from diabetic rats and age-matched controls at different time points: 1, 5, 10, and 15 weeks after diabetes modeling. (1)H nuclear magnetic resonance ((1)H NMR) spectra of the urine samples were obtained and analyzed by multivariate data analysis and quantitative statistical analysis. The metabolic patterns of diabetic groups are separated from the controls at each time point, suggesting that the metabolic profiles of diabetic rats were markedly different from the controls. Moreover, the samples from the diabetic 1-wk group are closely associated, whereas those of the diabetic 15-wk group are scattered, suggesting that the presence of various of complications contributes significantly to the pathogenesis of diabetes. Quantitative analysis indicated that urinary metabolites related to energy metabolism, tricarboxylic acid (TCA) cycle, and methylamine metabolism are involved in the evolution of diabetes. CONCLUSIONS/SIGNIFICANCE: The results highlighted that the numbers of metabolic changes were related to diabetes progression, and the perturbed metabolites represent potential metabolic biomarkers and provide clues that can elucidate the mechanisms underlying the generation and development of diabetes as well as its complication.

Acceleration of diabetic-wound healing with PEGylated rhaFGF in healing-impaired streptozocin diabetic rats.

Molecular modification with polyethylene glycol (PEGylation) is an effective approach to improve protein biostability, in vivo lifetime and therapeutic potency. In the present study, the recombinant human acid fibroblast growth factor (rhaFGF) was site-selectively PEGylated with 20 kDa mPEG-butyraldehyde. Mono-PEGylated rhaFGF was purified to near homogeneity by Sephadex G 25-gel filtration followed by a Heparin Sepharose TM CL-6B affinity chromatography. PEGylated rhaFGF has less effect than the native rhaFGF on the stimulation of 3T3 cell proliferation in vitro; however, its relative thermal stability at normal physiological temperature and structural stability were significantly enhanced, and its half-life time in vivo was significantly extended. Then, the physiological function of PEGylated rhaFGF on diabetic-wound healing was evaluated in type 1 diabetic Sprague Dawley rats. The results showed that, compared with the group of animal treated with native rhaFGF, the group treated with PEGylated rhaFGF exhibited better therapeutic efficacy with shorter healing time, quicker tissue collagen generation, earlier and higher transforming growth factor (TGF)-ß expression, and dermal cell proliferation. In addition, in vivo analysis showed that both native and PEGylated rhaFGF were more effective in the wound healing in the diabetic group compared with the nondiabetic one. Taken together, these results suggest that PEGylation of rhaFGF could be a more effective approach to the pharmacological and therapeutic application of native rhaFGF.

Improved refractory wound healing with administration of acidic fibroblast growth factor in diabetic rats.

The aim of the present study is to investigate the effect and mechanism of acidic fibroblast growth factor (aFGF) on treating refractory wound of diabetic rats. SD rats were randomly divided into control group, diabetes group, and aFGF group. Ulcer skin tissues of three groups of rats were respectively collected on days 7 and 14 after establishment of ulcer model for biochemical test, pathological section and immunohistochemistry to comprehensively evaluate the treatment effect of aFGF on diabetic ulcer. The results showed that aFGF could significantly increase capillaries and fibroblast amounts of ulcer tissues, enhance the expression of TGF-ß and PCNA proliferation proteins, and thus improved diabetic ulcer tissues. The preliminary mechanism that aFGF helps to promote healing of diabetic ulcer is possibly associated with that aFGF stimulated ulcer skins to secrete TGF-ß and PCNA proteins and promoted proliferation of capillaries and fibroblasts.

1H NMR-based metabonomic analysis of metabolic changes in streptozotocin-induced diabetic rats.

Diabetes mellitus is a complex metabolic disorder characterized by chronic hyperglycemia, hypoinsulinemia, and ketosis. To access the biochemical process of diabetes, we applied quantitative (1)H NMR-based metabonomics to analyze urine, serum, and liver extracts from streptozotocin-induced diabetic rats. Principle component analysis (PCA) of (1)H NMR spectra disclosed metabolic pattern differences between diabetic and control rats, and identified the related metabolic changes. The PCA scores plot demonstrated that the diabetic group could be distinguished from the control group, indicating that the metabolic characteristics of the two groups were markedly different. Our work reveals the accumulation of triglycerides, fatty acids and acetoacetate in diabetic rats, and may provide an efficient, convenient way for evaluating the pathological state and biochemical process of diabetes mellitus.

Equilibrium solubility and dissolution rate of the lead phosphate chloropyromorphite.

Phosphate addition to lead-contaminated soils can immobilize lead in situ through the formation of lead phosphate minerals such as chloropyromorphite (Pb5(PO4)3Cl). The long-term stability of lead immobilized in lead phosphate precipitates depends on the equilibrium solubility and dissolution rates of the lead phosphate solids. The equilibrium solubility and dissolution kinetics of chloropyromorphite were quantified in a series of batch and flow-through reactors. Both equilibrium solubility and dissolution rates were strongly affected by pH. Synthetic chloropyromorphite was more soluble than was predicted using a widely cited solubility product for pure chloropyromorphite, an observation that is consistent with several other recent studies. A trace amount of a more soluble lead solid, such as lead hydroxide, in chloropyromorphite could significantly increase dissolved Pb at neutral pH. The pH-dependence of the dissolution rate was examined in flow-through experiments. A dissolution rate law with a single rate constant for synthetic chloropyromorphite and a constant reaction order of 0.65 with respect to [H+] has been determined. The slow dissolution rate of chloropyromorphite relative to other lead minerals and its low solubility can result in its long-term stability in soils.