In vivo deep brain multiphoton fluorescence imaging emitting at NIR-I and NIR-II and excited at NIR-IV.

Multiphoton microscopy (MPM) enables deep brain imaging. Three optical windows: NIR-I, NIR-II, and NIR-III are widely used. Recently, NIR-IV (the 2200 nm window) has been demonstrated to be the last and longest window for deep tissue MPM. However, so far MPM covers only two optical windows labeled by single fluorescent probe, one for emission and one for excitation. Here we demonstrate in vivo deep brain MPM covering three optical windows, with emission at NIR-I, NIR-II, and excitation at NIR-IV, labeled by ICG. The innovations include: (1) characterizing both 3-photon excitation and emission properties of ICG emitting at both NIR-I and NIR-II, in water, plasma, and circulating blood; (2) a home-built multiphoton microscope with simultaneous dual channel detection, with which we demonstrate deep brain MPM 950 µm (NIR-I) and 850 µm (NIR-II) into the mouse brain in vivo, verifying that multi-optical window MPM is promising for deep brain imaging.

Comparison of the penetration depth in mouse brain in vivo through 3PF imaging using AIE nanoparticle labeling and THG imaging within the 1700 nm window.

3-Photon microscopy (3PM) excited at the 1700 nm window features a smaller tissue attenuation and hence a larger penetration depth in brain imaging compared with other excitation wavelengths in vivo. While the comparison of the penetration depth quantified by effective attenuation length le with other excitation wavelengths have been extensively investigated, comparison within the 1700 nm window has never been demonstrated. This is mainly due to the lack of a proper excitation laser source and characterization of the in vivo emission properties of fluorescent labels within this window. Herein, we demonstrate detailed measurements and comparison of le through the 3-photon imaging of the mouse brain in vivo, at different excitation wavelengths (1600 nm, 1700 nm, and 1800 nm). 3PF imaging and in vivo spectrum measurements were performed using AIE nanoparticle labeling. Our results show that le derived from both 3PF imaging and THG imaging is the largest at 1700 nm, indicating that it enables the deepest penetration in brain imaging in vivo.

In Vivo Deep-Brain 3- and 4-Photon Fluorescence Imaging of Subcortical Structures Labeled by Quantum Dots Excited at the 2200 nm Window.

Multiphoton microscopy (MPM) is an enabling technology for visualizing deep-brain structures at high spatial resolution in vivo. Within the low tissue absorption window, shifting to longer excitation wavelengths reduces tissue scattering and boosts penetration depth. Recently, the 2200 nm excitation window has emerged as the last and longest window suitable for deep-brain MPM. However, multiphoton fluorescence imaging at this window has not been demonstrated, due to the lack of characterization of multiphoton properties of fluorescent labels. Here we demonstrate technologies for measuring both the multiphoton excitation and emission properties of fluorescent labels at the 2200 nm window, using (1) 3-photon (ησ3) and 4-photon action cross sections (ησ4) and (2) 3-photon and 4-photon emission spectra both ex vivo and in vivo of quantum dots. Our results show that quantum dots have exceptionally large ησ3 and ησ4 for efficient generation of multiphoton fluorescence. Besides, the 3-photon and 4-photon emission spectra of quantum dots are essentially identical to those of one-photon emission, which change negligibly subject to the local environment of circulating blood. Based on these characterization results, we further demonstrate deep-brain vasculature imaging in vivo. Due to the superb multiphoton properties of quantum dots, 3-photon and 4-photon fluorescence imaging reaches a maximum brain imaging depth of 1060 and 940 µm below the surface of a mouse brain, respectively, which enables the imaging of subcortical structures. We thus fill the last gap in multiphoton fluorescence imaging in terms of wavelength selection.

Resolving arteriolar wall structures in mouse brain in vivo with three-photon microscopy.

The brain arteriolar wall is a multilayered structure, whose integrity is of key significance to the brain function. However, resolving these different layers in anmial models in vivo is hampered by the lack of either labeling or imaging technology. Here, we demonstrate that three-photon microscopy (3PM) is an ideal solution. In mouse brain in vivo, excited at the 1700-nm window, label-free third-harmonic generation imaging and three-photon fluorescence (3PF) imaging with Alexa 633 labeling colocalize and resolve the internal elastic lamina. Furthermore, Alexa Fluor 594-conjugated Wheat Germ Agglutinin (WGA-594) shows time-dependent labeling behavior. As time lapses, WGA-594 first labels endothelium, and then vascular smooth muscle cells, which are readily captured and resolved with 3PF imaging. Our results show that 3PM, in combination with proper labeling, is a promising technology for investigating the structures of brain arteriolar wall in vivo.

An effective preselection criterion for MODY with an increasingly positive genetic testing rate by NGS: results from two cohorts of Chinese children.

The purpose of this study was to determine the frequency of maturity-onset diabetes of the young (MODY) in two selected cohorts of Chinese children with diabetes and clinically suspected MODY, using next-generation sequencing (NGS). Ninety-three children who met the comprehensive criteria of suspected MODY were enrolled in two cohorts. A custom NGS panel or a whole exon group was used for sequencing. We identified 55/93 (59.1%) children with pathogenic and likely pathogenic MODY variants. Forty-two (76.3%) were confirmed to have the GCK (MODY2) mutation. Additionally, five had the HNF1A (MODY3), two the HNF1B (MODY5), one the 17q12 microdeletion (MODY5), two the HNF4A (MODY1), two the ABCC8 (MODY12), and one the PDX1 mutation (MODY4). Of these, 13 novel variants were detected in different genes. By comparing the gene-positive with gene-negative children, we found that discriminatory factors for MODY at diagnosis included lower HbA1c [7.4% vs. 10.2% (53 vs. 86 mmol/mol); P = 0.002], lower body mass index z score (0.2 vs. 1.0; P = 0.01), lower onset age (8.1 vs. 11.2 years; P = 0.001), and lower C-peptide (1.4 vs. 2.5 ng/mL; P = 0.02). In conclusion, the criteria used in this study for screening MODY are effective, and MODY2 is the most common subtype (76%), followed by MODY3 and MODY5. Some rare MODY subtypes have been reported in Chinese children.NEW & NOTEWORTHY We proved the clinical suspicion of maturity-onset diabetes of the young (MODY) according to the comprehensive criterion for next-generation sequencing testing, which helps to identify both common and rare MODYs, leading to accurate diagnosis and personalized treatment.

Analysis of the Incidence and Risk Factors of Precocious Puberty in Girls during the COVID-19 Pandemic.

Home quarantine due to the global coronavirus disease 2019 (COVID-19) pandemic has had a significant impact on children. Lifestyle changes have led to an increase in precocious puberty (PP) among girls, and the underlying risk factors for this remain unclear. Thus, we aimed to assess the influence of environmental, genetic, nutritional, and other lifestyle factors on the risk of PP in girls. We evaluated the incidence of new-onset PP in girls during home quarantine for COVID-19 and analyzed the potential risk factors. This was a retrospective questionnaire and medical record-based study involving 22 representative medical units from 13 cities in Henan Province, China. Girls with new-onset PP (central precocious puberty, 58; premature thelarche, 58; age, 5-9 years) between February 2020 and May 2020 were included, along with 124 healthy, age-matched controls. The number of new-onset PP cases reported during the study period was compared with that reported between February and May in 2018 and 2019. Patients' families completed a questionnaire to assess potential risk factors. There was a 5.01- and 3.14-fold increase in the number of new-onset PP cases from 2018 to 2020 and from 2019 to 2020, respectively; the differences were statistically significant (p < 0.01). High-risk factors for PP included longer time spent using electronic devices, decreased exercise time, higher body mass index, vitamin D deficiency, young age (<12 years) of mother during menarche, consumption of fried food and processed meat, residence in rural areas, and consumption of off-season fruits. Thus, we found that lifestyle changes caused due to the COVID-19 pandemic led to a significant increase in PP in girls. Management of the risk factors identified in this study may help in PP prevention.

Historical and recent change in extreme climate over East Asia.

This is an extended editors' commentary on the topical collection "Historical and recent change in extreme climate over East Asia", which collects a total of 15 papers related to the change and variability of extreme climate events in East Asia over the last few hundreds years. The extreme climate events are broadly classified into three categories: temperature and extreme warmth/coldness, precipitation and floods/droughts and western North Pacific typhoons. This commentary briefly summarizes the main findings presented in each paper in this topical collection, and outlines the implications of these findings for monitoring, detecting and modeling of regional climate change and for studying climate change impacts and adaptability. It also assesses the uncertainties of these studies, as well as the remaining knowledge gaps that should be filled in the future. One solid conclusion we can draw from these studies is that there was a marked decadal to multi-decadal variability of extreme climate events in East Asia in recent history, and the extreme events as observed during the last decades of the instrumental era were still within the range of natural variability except for some of those related to temperature. More severe and enduring droughts occurred in the early 20 th century or the earlier periods of history, frequently leading to great famines in northern China. Uncertainties remain in reconstructing historical extreme climate events and analyzing the early instrumental records. Further research could focus on the improvement of methodology in proxy based reconstruction of multi-decadal variations of surface air temperature and precipitation/drought, the recovery, digitization, calibration and verification of the early instrumental records, and the mechanisms of the observed multi-decadal variability of extreme climate in the region.

[Analysis of ALPL gene variant in a patient with infantile hypophosphatasia].

OBJECTIVE: To explore the genetic basis for a girl featuring bone and tooth mineralization disorder, premature deciduous teeth, rickets and short stature. METHODS: Genomic DNA was extracted and subjected to high-throughput whole exome sequencing. Suspected variants were confirmed by Sanger sequencing. Impact of potential variants was analyzed with bioinformatic software. RESULTS: The child was found to carry compound heterozygous missense variants of the ALPL gene, including c.1130C>T (p.A377V), a known pathogenic mutation inherited from her father, and c.1300G>A (p.V434M) inherited from her mother, which was unreported previously and predicted to be likely pathogenic based on standards and guidelines from the American College of Medical Genetics and Genomics (PM2+PM5+PP3+PP4). CONCLUSION: The compound heterozygous variants of c.1130C>T (p.Ala377Val) and c.1300G>A (p.Val434Met) of the ALPL gene probably underlay the disease in this child. Above finding has enriched the spectrum of ALPL gene variants.

Impact of the pandemic of COVID-19 on emergency attendance for stroke and acute myocardial infarction in Beijing, China.

To estimate the impact on emergency attendance for stroke and acute myocardial infarction (AMI) during the pandemic of COVID-19 in Beijing, China. Based on 17,123 and 8693 emergency attendance for stroke and AMI, an interrupted time-series (ITS) study was conducted. Since 01/24/2020, the top two levels of regulations on major public health have been implemented in Beijing. This study covered from 03/01/2018 to 06/03/2020, including 19 weeks of lockdown period and 99 weeks before. A segmented Poisson regression model was used to estimate the immediate change and the monthly change in the secular trend of the emergency attendance rates. The emergency attendance rates of stroke and AMI cut in half at the beginning of the lockdown period, with 52.1% (95% CI 45.8% to 57.7%) and 63.1% (95% CI 56.1% to 63.1%) immediate decreases for stroke and AMI, respectively. Then during the lockdown period, 7.0% (95% CI 2.5%, 11.6%) and 16.1% (95% CI 9.5, 23.1) increases per month in the secular trends of emergency attendance rates were shown for stroke and AMI, respectively. Though the accelerated increasing rates, there were estimated 1335 and 747 patients with stroke and AMI without seeking emergency medical aid during the lockdown, respectively. The emergency attendance for stroke and AMI cut in half at the beginning of the pandemic then had gradual restoration thereafter. The results hint the need for more engagement and communications with all stakeholders to reduce the negative impact on CVD emergency medical services during the crisis.

Sirolimus Therapy and Follow-up in a Patient with Severe Congenital Hyperinsulinism Following Subtotal Pancreatectomy

Congenital hyperinsulinism (CHI) is the most common cause of severe, persistent hypoglycemia in neonates and infants. If the patient does not respond to medical treatment the currently available treatment is subtotal pancreatectomy, but some patients still experience severe hypoglycemia after surgery. Sirolimus, a mammalian target of rapamycin inhibitor has recently been reported to be effective in the treatment of insulinoma and CHI patients. Here we report a patient with CHI who had prolonged hypoglycemia after subtotal pancreatectomy. The patient had a heterozygous mutation in ABCC8 but was unresponsive to an optimal dose of diazoxide (15 mg/ kg/day) and octreotide (30 µg/kg/day). The patient subsequently had subtotal pancreatectomy but severe and persistent hypoglycemia continued post-operatively. Sirolimus was commenced. There was a remarkable improvement in glycemic control without major adverse events, although he required a small dose of octreotide to maintain euglycemia. Sirolimus therapy was discontinued when the patient was 15 months old. At the time of this report, at an age of three years and eight months, the patient continues to maintain good glycemic control. This report suggests that sirolimus may be an effective treatment option in patients with CHI resistant to established medical therapy or failure of ubtotal pancreatectomy. However, the long-term safety requires study in larger groups of very young patients.

Immunosuppressive Siglec-E ligands on mouse aorta are up-regulated by LPS via NF-κB pathway.

AIMS: Siglec-E, the mouse ortholog of human Siglec-9, is an immunosuppressive cell surface receptor. Both Siglec-E and Siglec-9 are primarily found on neutrophils, macrophages, and monocytes. When Siglec-E binds to sialoglycan ligands in its extracellular environment, it halts the immune cells' inflammatory responses. In the present study, we aimed to investigate expression, mechanisms of action and regulation of Siglec-E ligands during vascular inflammation induced by E. coli lipopolysaccharides (LPS) in mouse aorta. METHODS: The distribution, molecular size and glycoprotein class of Siglec-E ligands on mouse aorta were determined, and the protein carrier of the ligands was identified. In vivo, the expression of Siglec-E ligands was detected after LPS treatment, with or without NF-κB inhibitor administration. In vitro, cultured primary mouse aortic endothelial cells (MAECs) were used to study changes in expression of Siglec-E ligands induced by LPS with or without NF-κB inhibitors. MAECs induced by LPS were co-cultured with macrophages and the effect of increased expression of Siglec-E ligands analyzed. RESULTS: Siglec-E ligands are O-linked sialoglycoproteins with molecular weights of 70-300 kDa and are distributed broadly on mouse aorta as well as on MAECs in vitro. In vivo, the expression of Siglec-E ligands was increased in mice aortas in response to LPS treatment in an NF-κB signaling pathway dependent manner. In MAECs, the expression of Siglec-E ligands was also increased by LPS via an NF-κB signaling pathway. Deleted in malignant brain tumors-1 was identified to be one of multiple protein carriers of Siglec-E ligands, and glycans of ligands involved in MAECs induced by LPS. Notably, co-incubation of macrophages with LPS-treated MAECs induced macrophage apoptosis and decreased macrophage phagocytosis, effects that were completely reversed by blocking Siglec-E binding to Siglec-E ligands. CONCLUSIONS: These data demonstrated that Siglec-E ligands were highly expressed in response to LPS-induced vascular inflammation and inhibited the immune response of macrophages, which may be a therapeutic strategy to interfere with vascular inflammation.

[Mutation analysis of a pedigree affected with brachydactyly type E2 and obesity].

OBJECTIVE: To identify pathogenic mutation in a pedigree affected with brachydactyly and obesity. METHODS: Peripheral blood sample was collected for extraction of genomic DNA. Exons capture combined with next generation sequencing (NGS) was carried out to identify potential mutation. Sanger sequencing was used to verify the results. RESULTS: NGS has identified a novel heterozygous missense mutation (c.125A>C, p.Gln42Pro) in the exon 1 of PTHLH gene. The result was verified by Sanger sequencing. The mutations was derived from his mother. His uncle and sister have also carried the same heterozygous mutation. CONCLUSION: A novel mutation of the PTHLH gene has been identified in a pedigree affected with brachydactyly type E2 and obesity.

Dysregulation of miR-135a-5p promotes the development of rat pulmonary arterial hypertension in vivo and in vitro.

Pulmonary arterial hypertension (PAH) is the most common form of pulmonary hypertension. Pulmonary arterial remodeling is closely related to the abnormal proliferation of pulmonary artery smooth muscle cells (PASMCs), which leads to the thickening of the medial layer of muscular arteries and then results in the narrowing or occlusion of the precapillary arterioles and PAH. However, the mechanisms underlying the abnormal proliferation of PASMCs remain unclear. In this study, we established rat primary PAH models using monocrotaline (MCT) injection or hypoxic exposure, then investigated the expression patterns of seven miRNAs associated with multiple pathogenic pathways central to pulmonary hypertension, and further explored the roles and the possible mechanisms of miR-135a during the development of PAH. In the rat primary PAH models, we observed that the expression of miR-135a-5p in lungs was drastically decreased at the initial stage of PAH development after MCT administration or hypoxic exposure, but it increased by 12-fold or 10-fold at the later stage. In vitro study in PASMCs showed a similar pattern of miR-135a-5p expression, with downregulation at 6 h but upregulation at 18, 24, and 48 h after hypoxic exposure. Early, but not late, administration of a miR-135a-5p mimic inhibited hypoxia-induced proliferation of PASMCs. The protective role of early miR-135a-5p agomir in the PAH rat model further supported the hypothesis that the early decrease in the expression of miR-135a-5p contributes to the proliferation of PASMCs and development of PAH, as early administration of miR-135a-5p agomir (10 nM, i.v.) reversed the elevated mean pulmonary arterial pressure and pulmonary vascular remodeling in MCT-treated rats. We revealed that miR-135a-5p directly bound to the 3'-UTR sequence of rat transient receptor potential channel 1 (TRPC1) mRNA and decreased TRPC1 protein expression, thus inhibiting PASMC proliferation. Collectively, our data suggest that dysregulation of miR-135a-5p in PASMCs contributes to the abnormal proliferation of PASMCs and the pathogenesis of PAH. Increasing miR-135a-5p expression at the early stage of PAH is a potential new avenue to prevent PAH development.

Immunoregulatory Siglec ligands are abundant in human and mouse aorta and are up-regulated by high glucose.

AIM: Inflammation is a driving force in development of atherosclerosis, and hyperglycemia is a significant risk factor for angiopathy. Siglec-9, expressed on human neutrophils and macrophages, engages specific glycan ligands on tissues to diminish ongoing inflammation. MATERIALS AND METHOD: Siglec-9 ligands on human aorta were characterized and the effects of high glucose exposure on the expression of ligands for Siglec-9 on human umbilical vein endothelial cells (HUV-EC-C) in vitro and ligands for the comparable siglec (Siglec-E) on mouse aorta in vivo were studied. KEY FINDINGS: Siglec-9 ligands were expressed broadly on human aorta, as well as on HUV-EC-C. Siglec-9 ligands on HUV-EC-C were sharply up-regulated under high glucose exposure in vitro, as were Siglec-E ligands on the aortas of hyperglycemic mice. Exposure of HUV-EC-C to high-glucose resulted in consistent inhibitory changes in co-cultured macrophages including increased apoptosis and decreased phagocytosis. Control of Siglec-9 ligand expression on HUV-EC-C was downstream of changes in an enzyme involved in their biosynthesis, UDP-galactose-4-epimerase (GALE) and increased cellular N-acetylgalactosamine. The alteration of GALE was associated with the regulatory microRNA hsa-let-7f. SIGNIFICANCE: We conclude that exposure to high-glucose results in up-regulation of immune inhibitory Siglec-9 sialoglycan ligands on aorta and HUV-EC-C cells downstream of altered GALE and GalNAc expression, resulting in up-regulation of apoptosis and decrease of phagocytic activity of macrophages. Changes in Siglec-9 sialoglycan ligand expression on vascular endothelial cells may be a natural response to the initial steps of atherosclerosis and might be a potential target to regulate inflammation in diabetic angiopathy.

[Protective effect of vitamin A on residual pancreatic ß cell function in children with type 1 diabetes mellitus].

OBJECTIVE: To study the protective effect of vitamin A on residual pancreatic ß cell function in children with type 1 diabetes mellitus (T1DM) and its mechanism. METHODS: A total of 46 children with T1DM (with a course of disease of 0.5-1 year) were randomly divided into an intervention group and a non-intervention group (n=23 each). The children in both groups were given insulin treatment, and those in the intervention group were also given vitamin A at a daily dose of 1 500-2 000 IU. A total of 25 healthy children were enrolled as the control group. The daily dose of insulin was calculated for the children with T1DM, and the serum levels of glycosylated hemoglobin (HbA1C), stimulated C-peptide, vitamin A, and interleukin-17 (IL-17) were measured before intervention and 3 months after intervention. RESULTS: Before vitamin A intervention, the intervention group and the non-intervention group had a significantly lower serum level of vitamin A and a significantly higher level of IL-17 than the control group (P<0.01). After 3 months of intervention, the intervention group had significantly lower serum IL-17 level and insulin dose and a significantly higher level of stimulated C-peptide than the non-intervention group (P<0.05). CONCLUSIONS: Vitamin A may protect residual pancreatic ß cell function, possibly by improving the abnormal secretion of IL-17 in children with T1DM.

Synthesis and Biological Evaluation of Ginsenoside Compound K Derivatives as a Novel Class of LXRα Activator.

Compound K is one of the active metabolites of Panaxnotoginseng saponins, which could attenuate the formation of atherosclerosis in mice modelsvia activating LXRα. We synthesized and evaluated a series of ginsenoside compound K derivatives modified with short chain fatty acids. All of the structures of this class of ginsenoside compound K derivative exhibited comparable or better biological activity than ginsenoside compound K. Especially structure 1 exhibited the best potency (cholesteryl ester content: 41.51%; expression of ABCA1 mRNA: 319%) and low cytotoxicity.

Genetic Analysis and Follow-Up of 25 Neonatal Diabetes Mellitus Patients in China.

AIMS: To study the clinical features, genetic etiology, and the correlation between phenotype and genotype of neonatal diabetes mellitus (NDM) in Chinese patients. METHODS: We reviewed the medical records of 25 NDM patients along with their follow-up details. Molecular genetic analysis was performed. We compared the HbA1c levels between PNDM group and infantile-onset T1DM patients. RESULTS: Of 25 NDM patients, 18 (72.0%) were PNDM and 7 (28.0%) were TNDM. Among 18 PNDM cases, 6 (33.3%) had known KATP channel mutations (KATP-PNDM). There were six non-KATP mutations, five novel mutations, including INS, EIF2AK3 (n = 2), GLIS3, and SLC19A2, one known EIF2AK3 mutation. There are two ABCC8 mutations in TNDM cases and one paternal UPD6q24. Five of the six KATP-PNDM patients were tried for glyburide transition, and 3 were successfully switched to glyburide. Mean HbA1c of PNDM was not significantly different from infantile onset T1DM (7.2% versus 7.4%, P = 0.41). CONCLUSION: PNDM accounted for 72% of NDM patients. About one-third of PNDM and TNDM patients had KATP mutations. The genetic etiology could be determined in 50% of PNDM and 43% of TNDM cases. PNDM patients achieved good glycemic control with insulin or glyburide therapy. The etiology of NDM suggests polygenic inheritance.

A parallel decoding algorithm for short polar codes based on error checking and correcting.

We propose a parallel decoding algorithm based on error checking and correcting to improve the performance of the short polar codes. In order to enhance the error-correcting capacity of the decoding algorithm, we first derive the error-checking equations generated on the basis of the frozen nodes, and then we introduce the method to check the errors in the input nodes of the decoder by the solutions of these equations. In order to further correct those checked errors, we adopt the method of modifying the probability messages of the error nodes with constant values according to the maximization principle. Due to the existence of multiple solutions of the error-checking equations, we formulate a CRC-aided optimization problem of finding the optimal solution with three different target functions, so as to improve the accuracy of error checking. Besides, in order to increase the throughput of decoding, we use a parallel method based on the decoding tree to calculate probability messages of all the nodes in the decoder. Numerical results show that the proposed decoding algorithm achieves better performance than that of some existing decoding algorithms with the same code length.