Laser-driven noncontact bubble transfer printing via a hydrogel composite stamp.

Transfer printing that enables heterogeneous integration of materials into spatially organized, functional arrangements is essential for developing unconventional electronic systems. Here, we report a laser-driven noncontact bubble transfer printing via a hydrogel composite stamp, which features a circular reservoir filled with hydrogel inside a stamp body and encapsulated by a laser absorption layer and an adhesion layer. This composite structure of stamp provides a reversible thermal controlled adhesion in a rapid manner through the liquid-gas phase transition of water in the hydrogel. The ultrasoft nature of hydrogel minimizes the influence of preload on the pick-up performance, which offers a strong interfacial adhesion under a small preload for a reliable damage-free pick-up. The strong light-matter interaction at the interface induces a liquid-gas phase transition to form a bulge on the stamp surface, which eliminates the interfacial adhesion for a successful noncontact printing. Demonstrations of noncontact transfer printing of microscale Si platelets onto various challenging nonadhesive surfaces (e.g., glass, key, wrench, steel sphere, dry petal, droplet) in two-dimensional or three-dimensional layouts illustrate the unusual capabilities for deterministic assembly to develop unconventional electronic systems such as flexible inorganic electronics, curved electronics, and micro-LED display.

scDecouple: decoupling cellular response from infected proportion bias in scCRISPR-seq.

Single-cell clustered regularly interspaced short palindromic repeats-sequencing (scCRISPR-seq) is an emerging high-throughput CRISPR screening technology where the true cellular response to perturbation is coupled with infected proportion bias of guide RNAs (gRNAs) across different cell clusters. The mixing of these effects introduces noise into scCRISPR-seq data analysis and thus obstacles to relevant studies. We developed scDecouple to decouple true cellular response of perturbation from the influence of infected proportion bias. scDecouple first models the distribution of gene expression profiles in perturbed cells and then iteratively finds the maximum likelihood of cell cluster proportions as well as the cellular response for each gRNA. We demonstrated its performance in a series of simulation experiments. By applying scDecouple to real scCRISPR-seq data, we found that scDecouple enhances the identification of biologically perturbation-related genes. scDecouple can benefit scCRISPR-seq data analysis, especially in the case of heterogeneous samples or complex gRNA libraries.

Resolution of Acinar Dedifferentiation Regulates Tissue Remodeling in Pancreatic Injury and Cancer Initiation.

BACKGROUND & AIMS: Acinar-to-ductal metaplasia (ADM) is crucial in the development of pancreatic ductal adenocarcinoma. However, our understanding of the induction and resolution of ADM remains limited. We conducted comparative transcriptome analyses to identify conserved mechanisms of ADM in mouse and human. METHODS: We identified Sox4 among the top up-regulated genes. We validated the analysis by RNA in situ hybridization. We performed experiments in mice with acinar-specific deletion of Sox4 (Ptf1a: CreER; Rosa26-LSL-YFPLSL-YFP; Sox4fl/fl) with and without an activating mutation in Kras (KrasLSL-G12D/+). Mice were given caerulein to induce pancreatitis. We performed phenotypic analysis by immunohistochemistry, tissue decellularization, and single-cell RNA sequencing. RESULTS: We demonstrated that Sox4 is reactivated in ADM and pancreatic intraepithelial neoplasias. Contrary to findings in other tissues, Sox4 actually counteracts cellular dedifferentiation and helps maintain tissue homeostasis. Moreover, our investigations unveiled the indispensable role of Sox4 in the specification of mucin-producing cells and tuft-like cells from acinar cells. We identified Sox4-dependent non-cell-autonomous mechanisms regulating the stromal reaction during disease progression. Notably, Sox4-inferred targets are activated upon KRAS inactivation and tumor regression. CONCLUSIONS: Our results indicate that our transcriptome analysis can be used to investigate conserved mechanisms of tissue injury. We demonstrate that Sox4 restrains acinar dedifferentiation and is necessary for the specification of acinar-derived metaplastic cells in pancreatic injury and cancer initiation and is activated upon Kras ablation and tumor regression in mice. By uncovering novel potential strategies to promote tissue homeostasis, our findings offer new avenues for preventing the development of pancreatic ductal adenocarcinoma.

Unconventional Piezoelectricity of Two-Dimensional Materials Driven by the Hall Effect.

Piezoelectricity has been widely explored for nanoelectromechanical applications, yet its working modes are mainly limited in polar directions. Here we discover the intrinsic electro-mechanical response in crystal materials that is transverse to the conventional polarized direction, which is named unconventional piezoelectricity. A Hall-like mechanism is proposed to interpret unconventional piezoelectricity as charge polarization driven by a built-in electric field for systems with asymmetric Berry curvature distributions. Density functional theory simulations and statistical analyses justify such a mechanism and confirm that unconventional piezoelectricity is a general property for various two-dimensional materials with spin splitting or valley splitting. An empirical formula is derived to connect the conventional and unconventional piezoelectricity. The extended understanding of the piezoelectric tensor in quantum materials opens an opportunity for applications in multidirectional energy conversion, broadband operation, and multifunctional sensing.

Ap-2ß regulates cranial osteogenic potential via the activation of Wnt/ß-catenin signaling pathway.

The skull is a fundamental bone that protects the development of brain and consists of several bony elements, such as the frontal and parietal bones. Frontal bone exhibited superior in osteogenic potential and regeneration of cranial defects compared to parietal bone. However, how this regional difference is regulated remains largely unknown. In this study, we identified an Ap-2ß transcriptional factor with a higher expression in frontal bone, but its molecular function in osteoblasts needs to be elucidated. We found that Ap-2ß knockdown in preosteoblasts leads to reduced proliferation, increased cell death and impaired differentiation. Through RNA-seq analysis, we found that Ap-2ß influences multiple signaling pathways including the Wnt pathway, and overexpression of Ap-2ß showed increased nuclear ß-catenin and its target genes expressions in osteoblasts. Pharmacological activation of Wnt/ß-catenin signaling using LiCl treatment cannot rescue the reduced luciferase activities of the ß-catenin/TCF/LEF reporter in Ap-2ß knockdown preosteoblasts. Besides, transient expression of Ap-2ß via the lentivirus system could sufficiently rescue the inferior osteogenic potential in parietal osteoblasts, while Ap-2ß knockdown in frontal osteoblasts resulted in reduced osteoblast activity, reduced active ß-catenin and target genes expressions. Taken together, our data demonstrated that Ap-2ß modulates osteoblast proliferation and differentiation through the regulation of Wnt/ß-catenin signaling pathway and plays an important role in regulating regional osteogenic potential in frontal and parietal bone.

Genomic and cytogenetic analyses reveal satellite repeat signature in allotetraploid okra (Abelmoschus esculentus).

BACKGROUND: Satellite repeats are one of the most rapidly evolving components in eukaryotic genomes and play vital roles in genome regulation, genome evolution, and speciation. As a consequence, the composition, abundance and chromosome distribution of satellite repeats often exhibit variability across various species, genome, and even individual chromosomes. However, we know little about the satellite repeat evolution in allopolyploid genomes. RESULTS: In this study, we investigated the satellite repeat signature in five okra (Abelmoschus esculentus) accessions using genomic and cytogenetic methods. In each of the five accessions, we identified eight satellite repeats, which exhibited a significant level of intraspecific conservation. Through fluorescence in situ hybridization (FISH) experiments, we observed that the satellite repeats generated multiple signals and exhibited variations in copy number across chromosomes. Intriguingly, we found that five satellite repeats were interspersed with centromeric retrotransposons, signifying their involvement in centromeric satellite repeat identity. We confirmed subgenome-biased amplification patterns of these satellite repeats through existing genome assemblies or dual-color FISH, indicating their distinct dynamic evolution in the allotetraploid okra subgenome. Moreover, we observed the presence of multiple chromosomes harboring the 35 S rDNA loci, alongside another chromosomal pair carrying the 5 S rDNA loci in okra using FISH assay. Remarkably, the intensity of 35 S rDNA hybridization signals varied among chromosomes, with the signals predominantly localized within regions of relatively weak DAPI staining, associated with GC-rich heterochromatin regions. Finally, we observed a similar localization pattern between 35 S rDNA and three satellite repeats with high GC content and confirmed their origin in the intergenic spacer region of the 35 S rDNA. CONCLUSIONS: Our findings uncover a unique satellite repeat signature in the allotetraploid okra, contributing to our understanding of the composition, abundance, and chromosomal distribution of satellite repeats in allopolyploid genomes, further enriching our understanding of their evolutionary dynamics in complex allopolyploid genomes.

The GaKAN2, a KANADI transcription factor, modulates stem trichomes in Gossypium arboreum.

KEY MESSAGE: GaKAN2, a member of the KANADI family, was found to be widely expressed in the cotton tissues and regulates trichome development through complex pathways. Cotton trichomes are believed to be the defense barrier against insect pests. Cotton fiber and trichomes are single-cell epidermal extensions with shared regulatory mechanisms. Despite several studies underlying mechanism of trichome development remains elusive. The KANADI is one of the key transcription factors (TFs) family, regulating Arabidopsis trichomes growth. However, the function of KANADI genes in cotton remains unknown. In the current study genome-wide scanning, transcriptomic analysis, gene silencing, subcellular localization, and yeast two-hybrid techniques were employed to decipher the function of KANADI TFs family genes in cotton crop. A total of 7 GaKAN genes were found in the Gossypium arboreum. Transcriptomic data revealed that these genes were significantly expressed in stem and root. Moreover, GaKAN2 was widely expressed in other tissues also. Subsequently, we selected GaKAN2 to validate the function of KANADI genes. Silencing of GaKAN2 resulted in a 24.99% decrease in single-cell trichomes and an 11.33% reduction in internodal distance, indicating its potential role in regulating trichomes and plant growth. RNA-Seq analysis elucidated that GaSuS and GaERS were the downstream genes of GaKAN2. The transcriptional activation and similarity in silencing phenotype between GaKAN2 and GaERS suggested that GaKAN2 regulates trichomes development through GaERS. Moreover, KEGG analysis revealed that a significant number of genes were enriched in the biosynthesis of secondary metabolites and plant hormone signal transduction pathways, thereby suggesting that GaKAN2 regulates the stem trichomes and plant growth. The GFP subcellular localization and yeast transcriptional activation analysis elucidated that GaKAN2 was located in the nucleus and capable of regulating the transcription of downstream genes. This study elucidated the function and characteristics of the KANADI gene family in cotton, providing a fundamental basis for further research on GaKAN2 gene in cotton plant trichomes and plant developmental processes.

Autophagy core protein BECN1 is vital for spermatogenesis and male fertility in mice†.

Mammalian spermatogenesis is a highly complex multi-step biological process, and autophagy has been demonstrated to be involved in the process of spermatogenesis. Beclin-1/BECN1, a core autophagy factor, plays a critical role in many biological processes and diseases. However, its function in spermatogenesis remains largely unclear. In the present study, germ cell-specific Beclin 1 (Becn1) knockout mice were generated and were conducted to determine the role of Becn1 in spermatogenesis and fertility of mice. Results indicate that Becn1 deficiency leads to reduced sperm motility and quantity, partial failure of spermiation, actin network disruption, excessive residual cytoplasm, acrosome malformation, and aberrant mitochondrial accumulation of sperm, ultimately resulting in reduced fertility in male mice. Furthermore, inhibition of autophagy was observed in the testes of germ cell-specific Becn1 knockout mice, which may contribute to impaired spermiogenesis and reduced fertility. Collectively, our results reveal that Becn1 is essential for fertility and spermiogenesis in mice.

The Role of Heat-Induced Stress Granules in the Blood-Testis Barrier of Mice.

Stress granules (SGs) are membraneless ribonucleoprotein (RNP)-based cellular foci formed in response to stress, facilitating cell survival by protecting against damage. Mammalian spermatogenesis should be maintained below body temperature for proper development, indicating its vulnerability to heat stress (HS). In this study, biotin tracer permeability assays showed that the inhibition of heat-induced SG assembly in the testis by 4-8 mg/kg cycloheximide significantly increased the percentage of seminiferous tubules with a damaged blood-testis barrier (BTB). Western blot results additionally revealed that the suppression of heat-induced SG assembly in Sertoli cell line, TM4 cells, by RNA inference of G3bp1/2 aggravated the decline in the BTB-related proteins ZO-1, ß-Catenin and Claudin-11, indicating that SGs could protect the BTB against damage caused by HS. The protein components that associate with SGs in Sertoli cells were isolated by sequential centrifugation and immunoprecipitation, and were identified by liquid chromatography with tandem mass spectrometry. Gene Ontology and KEGG pathway enrichment analysis revealed that their corresponding genes were mainly involved in pathways related to proteasomes, nucleotide excision repair, mismatch repair, and DNA replication. Furthermore, a new SG component, the ubiquitin associated protein 2 (UBAP2), was found to translocate to SGs upon HS in TM4 cells by immunofluorescence. Moreover, SG assembly was significantly diminished after UBAP2 knockdown by RNA inference during HS, suggesting the important role of UBAP2 in SG assembly. In addition, UBAP2 knockdown reduced the expression of ZO-1, ß-Catenin and Claudin-11, which implied its potential role in the function of the BTB. Overall, our study demonstrated the role of SGs in maintaining BTB functions during HS and identified a new component implicated in SG formation in Sertoli cells. These findings not only offer novel insights into the biological functions of SGs and the molecular mechanism of low fertility in males in summer, but also potentially provide an experimental basis for male fertility therapies.

Polysaccharide from Atractylodes macrocephala Koidz Binding with Zinc Oxide Nanoparticles as a Novel Mucosal Immune Adjuvant for H9N2 Inactivated Vaccine.

H9N2 avian influenza poses a significant public health risk, necessitating effective vaccines for mass immunization. Oral inactivated vaccines offer advantages like the ease of administration, but their efficacy often requires enhancement through mucosal adjuvants. In a previous study, we established a novel complex of polysaccharide from Atractylodes macrocephala Koidz binding with zinc oxide nanoparticles (AMP-ZnONPs) and preliminarily demonstrated its immune-enhancing function. This work aimed to evaluate the efficacy of AMP-ZnONPs as adjuvants in an oral H9N2-inactivated vaccine and the vaccine's impact on intestinal mucosal immunity. In this study, mice were orally vaccinated on days 0 and 14 after adapting to the environment. AMP-ZnONPs significantly improved HI titers, the levels of specific IgG, IgG1 and IgG2a in serum and sIgA in intestinal lavage fluid; increased the number of B-1 and B-2 cells and dendritic cell populations; and enhanced the mRNA expression of intestinal homing factors and immune-related cytokines. Interestingly, AMP-ZnONPs were more likely to affect B-1 cells than B-2 cells. AMP-ZnONPs showed mucosal immune enhancement that was comparable to positive control (cholera toxin, CT), but not to the side effect of weight loss caused by CT. Compared to the whole-inactivated H9N2 virus (WIV) group, the WIV + AMP-ZnONP and WIV + CT groups exhibited opposite shifts in gut microbial abundance. AMP-ZnONPs serve as an effective and safe mucosal adjuvant for oral WIV, improving cellular, humoral and mucosal immunity and microbiota in the gastrointestinal tract, avoiding the related undesired effects of CT.

Use of consecutive transcutaneous oxygen measurement when assessing the need for revascularization and association with the outcomes of ischemic diabetic ulcers.

This study compared the ankle-brachial index (ABI) with transcutaneous oxygen pressure (TcPO2 ) in assessing peripheral vascular disease (PVD) prevalence in 100 diabetic foot ulcer (DFU) patients. Patients were categorized into vascular or nonvascular reconstruction groups and underwent both ABI and TcPO2 measurements four times over 6 months. Predictive validity for PVD diagnosis was analysed using the area under the receiver-operating characteristic curve (AUC). The study found TcPO2 to be a superior predictor of PVD than ABI. Among the DFU patients, 51 with abnormal TcPO2 values underwent vascular reconstruction. Only TcPO2 values showed significant pretreatment differences between the groups and increased post-reconstruction. These values declined over a 6-month follow-up, whereas ABI values rose. For those with end-stage renal disease (ESRD), TcPO2 values saw a sharp decrease within 3 months. Pre-reconstruction TcPO2 was notably lower in amputation patients versus limb salvage surgery patients. In conclusion, TcPO2 is more effective than ABI for evaluating ischemic limb perfusion and revascularization necessity. It should be prioritized as the primary follow-up tool, especially for ESRD patients.

[Extracellular Matrix Stiffness Induces Mitochondrial Morphological Heterogeneity via AMPK Activation]. / 细胞外基质硬度通过AMPK调控干细胞线粒体的形态异质性.

Objective: To investigate the mechanical responses of mitochondrial morphology to extracellular matrix stiffness in human mesenchymal stem cells (hMSCs) and the role of AMP-activated protein kinase (AMPK) in the regulation of mitochondrial mechanoresponses. Methods: Two polyacrylamide (PAAm) hydrogels, a soft one with a Young's modulus of 1 kPa and a stiff one of 20 kPa, were prepared by changing the monomer concentrations of acrylamide and bis-acrylamide. Then, hMSCs were cultured on the soft and stiff PAAm hydrogels and changes in mitochondrial morphology were observed using a laser confocal microscope. Western blot was performed to determine the expression and activation of AMPK, a protein associated with mitochondrial homeostasis. Furthermore, the activation of AMPK was regulated on the soft and stiff matrixes by AMPK activator A-769662 and the inhibitor Compound C, respectively, to observe the morphological changes of mitochondria. Results: The morphology of the mitochondria in hMSCs showed heterogeneity when there was a change in gel stiffness. On the 1 kPa soft matrix, 74% mitochondria exhibited a dense, elongated filamentous network structure, while on the 20 kPa stiff matrix, up to 63.3% mitochondria were fragmented or punctate and were sparsely distributed. Western blot results revealed that the phosphorylated AMPK (p-AMPK)/AMPK ratio on the stiff matrix was 1.6 times as high as that on the soft one. Immunofluorescence assay results revealed that the expression of p-AMPK was elevated on the hard matrix and showed nuclear localization, which indicated that the activation of intracellular AMPK increased continuously along with the increase in extracellular matrix stiffness. When the hMSCs on the soft matrix were treated with A-769662, an AMPK activator, the mitochondria transitioned from a filamentous network morphology to a fragmented morphology, with the ratio of filamentous network decreasing from 74% to 9.5%. Additionally, AMPK inhibition with Compound C promoted mitochondrial fusion on the stiff matrix and significantly reduced the generation of punctate mitochondria. Conclusion: Extracellular matrix stiffness regulates mitochondrial morphology in hMSCs through the activation of AMPK. Stiff matrix promotes the AMPK activation, resulting in mitochondrial fission and the subsequent fragmentation of mitochondria. The impact of matrix stiffness on mitochondrial morphology can be reversed by altering the level of AMPK phosphorylation.

miRNA-124 alleviated memory impairment induced by d-galactose rapidly in male rats via microglia polarization.

MiRNA-124 has been considered to play a significant role in the formation of memory and a variety of neurodegenerative diseases. In this study, the aim is to verify whether miRNA-124 is involved in memory impairment induced by d-galactose, and explore the underlying neuroprotective mechanism. The results revealed that rapid administration of d-galactose (1000 mg/kg subcutaneously) in mice caused memory impairments, as determined by Novel Object Recognition test, Morris Water Maze test, and histological assessments. MiRNA-124 agomir is stereotactic injected into hippocampus, thus alleviated memory impairment induced by d-galactose and reversed the neural damage and neuroinflammation. Furthermore, the results of molecular biological analysis and immunohistochemistry revealed that miRNA-124 markedly reduced neuroinflammation induced by d-galactose through polarization of microglia as determined by detection of ionized calcium binding adapter molecule 1 (Iba-1), inducible nitric oxide synthase (iNOS) and arginase-1(Arg-1), which also downregulated inflammatory mediators, including interleukin-1ß (IL-1ß) and tumor necrosis factor-α (TNF-α), and upregulated IL-4 and IL-10. Hence, taken together, the results of the present study suggested that miRNA-124 showed a significant negative correlation with memory impairment and neuroinflammation induced by d-galactose rapidly, possibly via polarization of microglia from M1 to M2. It is possible that miRNA-124 can be used as a new target for the pathogenesis of memory impairment, including age-associated neurodegenerative diseases such as Alzheimer's disease.

Characterization of stemness features and construction of a stemness subtype classifier to predict survival and treatment responses in lung squamous cell carcinoma.

BACKGROUND: Cancer stemness has been proven to affect tumorigenesis, metastasis, and drug resistance in various cancers, including lung squamous cell carcinoma (LUSC). We intended to develop a clinically applicable stemness subtype classifier that could assist physicians in predicting patient prognosis and treatment response. METHODS: This study collected RNA-seq data from TCGA and GEO databases to calculate transcriptional stemness indices (mRNAsi) using the one-class logistic regression machine learning algorithm. Unsupervised consensus clustering was conducted to identify a stemness-based classification. Immune infiltration analysis (ESTIMATE and ssGSEA algorithms) methods were used to investigate the immune infiltration status of different subtypes. Tumor Immune Dysfunction and Exclusion (TIDE) and Immunophenotype Score (IPS) were used to evaluate the immunotherapy response. The pRRophetic algorithm was used to estimate the efficiency of chemotherapeutic and targeted agents. Two machine learning algorithms (LASSO and RF) and multivariate logistic regression analysis were performed to construct a novel stemness-related classifier. RESULTS: We observed that patients in the high-mRNAsi group had a better prognosis than those in the low-mRNAsi group. Next, we identified 190 stemness-related differentially expressed genes (DEGs) that could categorize LUSC patients into two stemness subtypes. Patients in the stemness subtype B group with higher mRNAsi scores exhibited better overall survival (OS) than those in the stemness subtype A group. Immunotherapy prediction demonstrated that stemness subtype A has a better response to immune checkpoint inhibitors (ICIs). Furthermore, the drug response prediction indicated that stemness subtype A had a better response to chemotherapy but was more resistant to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs). Finally, we constructed a nine-gene-based classifier to predict patients' stemness subtype and validated it in independent GEO validation sets. The expression levels of these genes were also validated in clinical tumor specimens. CONCLUSION: The stemness-related classifier could serve as a potential prognostic and treatment predictor and assist physicians in selecting effective treatment strategies for patients with LUSC in clinical practice.

Dietary fiber modification: structure, physicochemical properties, bioactivities, and application-a review.

There is increasing attention on the modification of dietary fiber (DF), since its effective improvement on properties and functions of DF. Modification of DF can change their structure and functions to enhance their bioactivities, and endow them with huge application potential in the field of food and nutrition. Here, we classified and explained the different modification methods of DF, especially dietary polysaccharides. Different modification methods exert variable effects on the chemical structure of DF such as molecular weight, monosaccharide composition, functional groups, chain structure, and conformation. Moreover, we have discussed the change in physicochemical properties and biological activities of DF, resulting from alterations in the chemical structure of DF, along with a few applications of modified DF. Finally, we have summarized the modified effects of DF. This review will provide a foundation for further studies on DF modification and promote the future application of DF in food products.

Antibiotic-loaded bone substitutes therapy in the management of the moderate to severe diabetic foot infection: A meta-analysis.

In recent years, local antibiotic-loaded bone substitutes (ALBS) have been used increasingly in the treatment of diabetic foot infection (DFI). The meta-analysis aimed to analyse the efficacy of ALBS on patients with moderate to severe DFI (with or without osteomyelitis). With an appropriate search strategy, 7 studies were selected for analysis (2 RCTs and 5 cohort studies). The result showed that the application of ALBS effectively reduced the length of hospital stay (WMD -5.55; 95% CI: -9.85 to -1.26; P = 0.01), the recurrence rates (RR 0.33; 95% CI: 0.15 to 0.69; P = 0.003) and the mortality rates (RR 0.22; 95% CI: 0.06 to 0.82; P = 0.02). Compared to the control groups, however, there was no difference in healing rates (RR 1.06; 95% CI: 0.96 to 1.18; P = 0.26), healing time (WMD -1.44; 95% CI: -3.37 to -0.49; P = 0.14), the number of debridement (WMD -1.98; 95% CI: -4.08 to 0.12; P = 0.06) and major amputation rates (RR 0.76; 95% CI: 0.35 to 1.61; P = 0.47). The ALBS appears to have some beneficial effects as an adjunct to standard surgery in the treatment of DFI with or without osteomyelitis, as it reduces recurrence rates, mortality rates, and length of hospital stay, but there was no statistically significant difference in enhancing wound healing.

Prediction of nonsentinel lymph node metastasis in acral melanoma with positive sentinel lymph nodes.

BACKGROUND: Metastasis in a nonsentinel lymph node (non-SLN) is an unfavorable independent prognostic factor in cutaneous melanoma (CM). Recent data did suggest potential value of completion lymph node dissection (CLND) in CM patients with non-SLN metastasis. Prediction of non-SLN metastasis assists clinicians in deciding on adjuvant therapy without CLND. We analyzed risk factors and developed a prediction model for non-SLN status in acral melanoma (AM). METHODS: This retrospective study enrolled 656 cases of melanoma who underwent sentinel lymph node biopsy at Fudan University Shanghai Cancer Center from 2009 to 2017. We identified 81 SLN + AM patients who underwent CLND. Clinicopathologic data, including SLN tumor burden and non-SLN status were examined with Cox and Logistics regression models. RESULTS: Ulceration, Clark level, number of deposits in the SLN (NumDep) and maximum size of deposits (MaxSize) are independent risk factors associated with non-SLN metastases. We developed a scoring system that combines ulceration, the cutoff values of Clark level V, MaxSize of 2 mm, and NumDep of 5 to predict non-SLN metastasis with an efficiency of 85.2% and 100% positive predictive value in the high-rank group (scores of 17-24). CONCLUSIONS: A scoring system that included ulceration, Clark level, MaxSize, and NumDep is reliable and effective for predicting non-SLN metastasis in SLN-positive AM.

Assessment of fetal intraventricular diastolic fluid dynamics using ultrasound vector flow mapping.

OBJECTIVE: The purpose of this study was to investigate the feasibility of visualizing and quantifying the normal pattern of vortex formation in the left ventricle (LV) and right ventricle (RV) of the fetal heart during diastole using vector flow mapping (VFM). METHODS: A total of 36 healthy fetuses in the second trimester (mean gestational age: 23 weeks, 2 days; range: 22-24 weeks) were enrolled in the study. Color Doppler signals were recorded in the four-chamber view to observe the phase of the diastolic vortices in the LV and RV. The vortex area and circulation were measured, and parameters such as intraventricular pressure difference (IVPD), intraventricular pressure gradient (IVPG), and average energy loss (EL_AVG) were evaluated at different diastolic phases, including isovolumic relaxation (D1), early diastole (D2), and late diastole (D3). RESULTS: Healthy second-trimester fetal vortex formations were observed in both the LV and RV at the end of diastole, with the vortices rotating in a clockwise direction towards the outflow tract. There were no significant differences in vortex area and circulation between the two ventricles (p > 0.05). However, significant differences were found in IVPD, IVPG, and EL_AVG among the diastolic phases (D1, D2, and D3) (p < 0.05). Trends in IVPD, IVPG, and EL_AVG during diastole (D1-D2-D3) revealed increasing IVPD and EL_AVG values, as well as decreasing IVPG values. Furthermore, during D3, the RV exhibited significantly higher IVPD, IVPG, and EL_AVG compared to the LV (p > 0.05). CONCLUSION: VFM is a valuable technique for analyzing the formation of vortices in the left and right ventricles during fetal diastole. The application of VFM technology has the potential to enhance the assessment of fetal cardiac parameters.

Multiplexed Homogeneous Immunoassay Based on Counting Single Immunocomplexes together with Dark-Field and Fluorescence Microscopy.

The development of multiplexed immunoassays is impeded by the difficulty in distinguishing labeled immunocomplexes from free probes and nonspecifically bound probes. Here, we attempted to overcome this issue by counting core-satellite-structured immunocomplexes simultaneously using dark-field and fluorescence microscopy. The tumor biomarkers of carcinoembryonic antigen (CEA), α-fetoprotein (AFP), and prostate-specific antigen (PSA) were chosen as model targets. Gold nanoparticles (AuNPs) with diameters of 70 nm were coated with the detection antibodies of the three targets. Quantum dot (QD) 525, QD 585, and QD 655 were modified with the capture antibodies of CEA, AFP, and PSA, respectively. Then, an immunocomplex containing one AuNP and one or several QDs was formed, whereas free and nonspecifically bound probes had either one AuNP or one QD. When observed with a transmission grating-based spectral microscope, the immunocomplexes had overlapping scattering and fluorescent spectral images and were therefore identified and quantified precisely. The biomarkers inside the immunocomplexes were recognized on the basis of the fluorescent first-order streaks of the QDs. Model biomarkers in buffer and in 12.6% blank plasma were quantified for validation. The limits of detection for CEA, PSA, and AFP in buffer were in dozens of femtomolar and were close to those in blank plasma. The results demonstrated that our approach worked well in distinguishing immunocomplexes from free and nonspecifically bound probes. The successful quantification of the three targets in five human plasma samples verified the reliability of our method in clinical applications.

Deficiency of vitamin D receptor in keratinocytes augments dermal fibrosis and inflammation in a mouse model of HOCl-induced scleroderma.

Scleroderma, characterized by extensive fibrosis and vascular alterations, involves excessive fibroblast activation, uncontrolled inflammation, and abnormal collagen deposition. Previous studies showed that administrations of either 1,25(OH)2D3 or vitamin D analog effectively decreased or reversed skin fibrosis by regulating the extracellular matrix homeostasis. The actions of 1,25(OH)2D3 are mediated by the vitamin D receptor (VDR), a transcription regulator crucial for skin homeostasis. Although evidence suggests that keratinocyte-fibroblast interaction influences the development of scleroderma, the role of keratinocytes in scleroderma remains unknown. Here, we demonstrated that the ablation of VDR in keratinocytes greatly exacerbated dermal fibrosis in HOCl-induced scleroderma in mice. The deficiency of VDR in the epidermis marked increased dermal thickness, inflammatory cell infiltration, and severe collagen deposition in comparison to the control group in HOCl-treated skin. Moreover, significant elevations in expression levels of mRNA for collagen overproduction (Col1A1, Col1A2, Col3A1, α-SMA, MMP9, TGF-ß1) and proinflammatory cytokines (IL-1ß, IL-6, CXCL1, CXCL2) were observed in VDR conditional KO versus control mice following HOCl treatment. Collectively, these results suggest that VDR in keratinocytes plays a pivotal role in scleroderma progression, and the interplay between keratinocytes and fibroblasts deserves more attention regarding the exploration of the pathogenesis and treatment for scleroderma.