Inkjet Printing Photoresist with Ultralow Viscosity on Silicon Wafers for Uniform Coating.

Inkjet printing is introduced into the photoresist coating process for uniform photoresist film formation on silicon wafers with the in-house inkjet experimental prototype. The optimization of a dual negative voltage waveform is proposed to achieve stable droplet jetting for the ultralow viscosity (0.71 mPa·s) photoresist with a 1:10 dilution ratio employed in the semiconductor packaging processes. Moreover, the maximum droplet jetting velocity can reach 9.51 m/s, and the droplet volume is controlled at ∼6.5 pL with excellent droplet concentration. The uniform film of the AZ P4620 photoresist is coated on silicon wafers by quantitatively exploring and optimizing the printhead driving frequency and movement velocity utilizing the droplet deposition model and experimental analysis. Results show that the optimal inkjet parameters with 5 kHz in jetting frequency and 6 mm/s in motion velocity can obtain a film evenness index of 4.81% with the thickness of 0.945 µm, which exhibits a more uniform photoresist film than the spray coating method. The study not only expands the application of the inkjet printing technique but also offers an alternative for photoresist coating in the photolithography process.

In vivo pharmacodynamic study of contezolid acefosamil, a prodrug of contezolid for oral and intravenous administration.

OBJECTIVES: Contezolid acefosamil is a novel O-acyl phosphoramidate prodrug of contezolid. In the current study, we aimed to systemically evaluate the efficacy of contezolid acefosamil against infections caused by multiple Gram-positive pathogens, and compare the efficacy of the prodrug by oral and intravenous administrations. METHODS: The in vivo pharmacodynamic efficacy of contezolid acefosamil was evaluated in mouse models of systemic (with five S. aureus, three S. pneumoniae and two S. pyogenes bacterial isolates) and thigh (with two S. aureus isolates) infections using linezolid as the reference agent. RESULTS: In both models, contezolid acefosamil administrated either orally or intravenously, demonstrated high antibacterial efficacy similar to linezolid, and the antibacterial efficacy of oral and intravenous contezolid acefosamil were comparable. CONCLUSIONS: The high aqueous solubility and great efficacy of contezolid acefosamil support its clinical development as an injectable and oral antibiotic suitable for serious Gram-positive infections.

Bruceine A protects against diabetic kidney disease via inhibiting galectin-1.

Bruceine A is a natural quassinoid compound extracted from the fruit of the Traditional Chinese Medicine Brucea javanica (L.) Merr. that has various types of various biological activities. However, whether the compound has a protective effect on diabetic kidney disease remains unknown. Galectin-1 is actively involved in a variety of chronic inflammation-relevant human diseases including diabetic kidney disease. Here, we identified Bruceine A as a kidney protective molecule against a model of diabetic kidney disease in db/db mice with potent anti-inflammatory activity both in vitro and in vivo. Mechanistically, by selectively binding to the conserved carbohydrate-recognition domain of galectin-1 and disrupting the interaction between galectin-1 and the receptor for activated protein C kinase 1, Bruceine A was found to inhibit galectin-1-mediated inflammatory signal transduction under high glucose stress in rat mesangial HBZY-1 cells. Thus, our findings reveal Bruceine A as an unidentified galectin-1 inhibitor affording significant protection against diabetic kidney disease and may provide novel pharmacological therapeutics for the disease.

Activation of the sigma-1 receptor exerts cardioprotection in a rodent model of chronic heart failure by stimulation of angiogenesis.

BACKGROUND: Angiogenesis plays a critical role on post-infarction heart failure (PIHF), the presence of which facilitates additional blood supply to maintain the survival of residual cardiomyocytes. The sigma-1 receptor (S1R) has been substantiated to stimulate angiogenesis, with the effect on a model of PIHF remaining unknown. AIMS: This study aims to investigate the effects of S1R on PIHF and the underlying mechanisms involved. METHODS: Rats were implemented left anterior descending artery ligation followed by rearing for 6 weeks to induce a phenotype of heart failure. Daily intraperitoneal injection of S1R agonist or antagonist for 5 weeks was applied from 2nd week after surgery. The effects exerted by S1R were detected by echocardiography, hemodynamic testing, western blot, Sirius red dyeing, ELISA, immunohistochemistry and fluorescence. We also cultured HUVECs to verify the mechanisms in vitro. RESULTS: Stimulation of S1R significantly ameliorated the cardiac function resulted from PIHF, in addition to the observation of reduced fibrosis in the peri-infarct region and the apoptosis of residual cardiomyocytes, which were associated with augmentation of microvascular density in peri-infarct region through activation of the JAK2/STAT3 pathway. We also indicated that suppression of JAK2/STAT3 pathway by specific inhibitor in vitro reversed the pro-angiogenic effects of S1R on HUVECs, which further confirmed that angiogenesis, responsible for PIHF amelioration, by S1R stimulation was in a JAK2/STAT3 pathway-dependent manner. CONCLUSION: S1R stimulation improved PIHF-induced cardiac dysfunction and ventricular remodeling through promoting angiogenesis by activating the JAK2/STAT3 pathway.

PHF7 is a novel histone H2A E3 ligase prior to histone-to-protamine exchange during spermiogenesis.

Epigenetic regulation, including histone-to-protamine exchanges, controls spermiogenesis. However, the underlying mechanisms of this regulation are largely unknown. Here, we report that PHF7, a testis-specific PHD and RING finger domain-containing protein, is essential for histone-to-protamine exchange in mice. PHF7 is specifically expressed during spermiogenesis. PHF7 deletion results in male infertility due to aberrant histone retention and impaired protamine replacement in elongated spermatids. Mechanistically, PHF7 can simultaneously bind histone H2A and H3; its PHD domain, a histone code reader, can specifically bind H3K4me3/me2, and its RING domain, a histone writer, can ubiquitylate H2A. Thus, our study reveals that PHF7 is a novel E3 ligase that can specifically ubiquitylate H2A through binding H3K4me3/me2 prior to histone-to-protamine exchange.

Efficacy of Human-Recombinant Epidermal Growth Factor Combined with Povidone-Iodine for Pressure Ulcers and Its Influence on Inflammatory Cytokines.

Purpose: To determine the clinical efficacy of recombinant human epidermal growth factor (rh-EGF) combined with povidone-iodine (PVI) on patients with pressure ulcers (PUs). Methods: One hundred and five PU patients treated between January 2018 and January 2021 were enrolled and retrospectively analyzed. Of them, 50 patients who received conventional treatment were assigned to the control group (Con group), while 55 patients treated with rh-EGF combined with PVI were assigned to the observation group (Obs group). The two groups were compared in clinical efficacy, PU alleviation (total area reduction rate, total depth reduction rate, and total volume reduction rate), healing time, pain degree (Visual Analog Scale [VAS] score), inflammatory indexes (interleukin-8 [IL-8], tumor necrosis factor-α [TNF-α], and hypersensitive C reactive protein [hs-CRP]), and hydroxyproline content in the wound. Results: The Obs group yielded a higher total effective rate than the Con group (P < 0.05). The Obs group also experienced statistically shorter healing time and milder pain, with better PU alleviation and lower levels of inflammation indexes compared with the Con group (all P < 0.05). In addition, a higher hydroxyproline content in the wound was found in the Obs group. Conclusions: All in all, rh-EGF combined with PVI has a definite curative effect on patients with PUs. It can promote PU alleviation and hydroxyproline secretion in the wound and inhibit pain and inflammatory reactions, which is worthy of clinical promotion.

Chondroitin synthase-3 regulates nucleus pulposus degeneration through actin-induced YAP signaling.

Loss of chondroitin sulfate (CS) has been reported to play a key role during intervertebral disc degeneration (IDD). However, the detailed mechanism of CS and its synthases have not been elucidated. Since CS is mainly synthesized by chondroitin synthases 3 (Chsy3), here, the Chsy3 knockout mice are generated by using CRISPR-Cas9 and semi-cloning technology to study its mechanism during IDD. We find that CS and Chsy3 expression are decreased during IDD both in human and mice nucleus pulposus (NP) tissue, and knockout of Chsy3 shows that spontaneous IDD phenotype resembles that of human samples in the Chsy3-/- mice. Taking advantage of RNA-Seq data, we confirm increased catabolic and decreased anabolic changes in Chsy3-/- NP cells. By using bioinformatic analysis and validation, we find that Hippo signaling pathway is significantly downregulated, and the activation of Yap1 is mainly affected in Chsy3-/- NP cells. Furthermore, functional analyses have shown that Chsy3 could regulate NP cell degeneration by Actin tension mediated activation of Yap1, which is independent of Hippo/Lats signaling. In summary, our findings reveal a novel mechanism that depletion of CS-related Chsy3 can cause spontaneous intervertebral disc degeneration by mediating Yap activation through CS-related actin-tension in NP cells.

The combined antibacterial effects of sodium new houttuyfonate and berberine chloride against growing and persistent methicillin-resistant and vancomycin-intermediate Staphylococcus aureus.

BACKGROUND: Infections caused by drug-resistant Staphylococcus aureus, especially vancomycin-intermediate Staphylococcus aureus (VISA), leave clinicians with limited therapeutic options for treatment. Persister cells is a leading cause of recalcitrant infection and antibiotic treatment failure, and there is no drug in clinical use that specifically targets persister cells currently. Here, we report a promising combination therapy of sodium new houttuyfonate (SNH) and berberine chloride (BBR) which is able to eradicate both growing and persistent drug-resistant Staphylococcus aureus. RESULTS: The susceptibility test showed SNH exhibited anti-MRSA activity with MIC90 at 64 µg/mL, while BBR showed weak anti-MRSA activity with MIC90 at 512 µg/mL. MICs of BBR in combination with 1/2 MIC SNH decreased by 4 to 64 folds compared with MICs of BBR alone. The results of time-killing assays revealed that the combined use of sub-MIC SNH and BBR offered an in vitro synergistic action against growing MRSA (including pathogenic MRSA) and VISA strains. More importantly, the combination of SNH and BBR was able to eradicate VISA Mu50 and pathogenic MRSA persister cells. The synergistic effect is likely related to the interruption of the cell membrane caused by SNH, which is confirmed by scanning electron microscope and membrane potential and permeability analysis. CONCLUSIONS: Our study provide a promising clinical curative strategy for combating drug-resistant S. aureus infections, especially for recalcitrant infections caused by persister cells.

Synergistic Effect of Colistin Combined with PFK-158 against Colistin-Resistant Enterobacteriaceae.

As increasing numbers of colistin-resistant bacteria emerge, new therapies are urgently needed to treat infections caused by these pathogens. The discovery of new combination therapies is one important way to solve such problems. Here, we report that the antitumor drug PFK-158 and its analogs PFK-015 and 3PO can exert synergistic effects with colistin against colistin-resistant Enterobacteriaceae, including mcr-1-positive or high-level-colistin-resistant (HLCR) isolates, as shown by a checkerboard assay. The results of a time-kill assay revealed that colistin combined with PFK-158 continuously eliminated colistin-resistant Escherichia coli 13-43, Klebsiella pneumoniae H04, and Enterobacter cloacae D01 in 24 h. Images from scanning electron microscopy (SEM) at 5 h postinoculation confirmed the killing effect of the combination. Finally, in vivo treatment showed that PFK-158 had a better synergistic effect than its analogs. Compared to the corresponding rates after colistin monotherapy, the survival rates of systemically infected mice were significantly increased 30% or 60% when the mice received an intravenous injection of colistin in combination with 15 mg/kg of body weight PFK-158. These results have important implications for repurposing PFK-158 to combat colistin resistance.

Daphnetin: A Novel Anti-Helicobacter pylori Agent.

BACKGROUND: Antibiotic-resistant H. pylori was increasingly found in infected individuals, which resulted in treatment failure and required alternative therapeutic strategies. Daphnetin, a coumarin-derivative compound, has multiple pharmacological activities. METHODS: The mechanism of daphnetin on H. pylori was investigated focusing on its effect on cell morphologies, transcription of genes related to virulence, adhesion, and cytotoxicity to human gastric epithelial (GES-1) cell line. RESULTS: Daphnetin showed good activities against multidrug resistant (MDR) H. pylori clinical isolates, with minimal inhibitory concentration (MIC) values ranging from 25 to 100 µg/mL. In addition, daphnetin exposure resulted in H. pylori morphological changes. Moreover, daphnetin caused increased translocation of phosphatidylserine (PS), DNA damage, and recA expression, and RecA protein production vs. control group. Of great importance, daphnetin significantly decreased H. pylori adhesion to GES-1 cell line vs. control group, which may be related to the reduced expression of colonization related genes (e.g., babA and ureI). CONCLUSIONS: These results suggested that daphnetin has good activity against MDR H. pylori. The mechanism(s) of daphnetin against H. pylori were related to change of membrane structure, increase of DNA damage and PS translocation, and decrease of H. pylori attachment to GES-1 cells.

An intermediate cell state allows rerouting of cell fate.

Somatic cell reprogramming into pluripotent stem cells using transcriptional factors or chemical compounds has been shown to include an intermediate cell state with epithelial features. Two recent papers show that this intermediate state can be redirected to create other differentiated cell types-specifically hepatocytes and neurons-using chemical cocktails. These results shed new light on a critical intermediate in cell fate conversion with mechanistic and practical implications.

Nanoliter-Scale Oil-Air-Droplet Chip-Based Single Cell Proteomic Analysis.

Single cell proteomic analysis provides crucial information on cellular heterogeneity in biological systems. Herein, we describe a nanoliter-scale oil-air-droplet (OAD) chip for achieving multistep complex sample pretreatment and injection for single cell proteomic analysis in the shotgun mode. By using miniaturized stationary droplet microreaction and manipulation techniques, our system allows all sample pretreatment and injection procedures to be performed in a nanoliter-scale droplet with minimum sample loss and a high sample injection efficiency (>99%), thus substantially increasing the analytical sensitivity for single cell samples. We applied the present system in the proteomic analysis of 100 ± 10, 50 ± 5, 10, and 1 HeLa cell(s), and protein IDs of 1360, 612, 192, and 51 were identified, respectively. The OAD chip-based system was further applied in single mouse oocyte analysis, with 355 protein IDs identified at the single oocyte level, which demonstrated its special advantages of high enrichment of sequence coverage, hydrophobic proteins, and enzymatic digestion efficiency over the traditional in-tube system.

An HDAC2-TET1 switch at distinct chromatin regions significantly promotes the maturation of pre-iPS to iPS cells.

The maturation of induced pluripotent stem cells (iPS) is one of the limiting steps of somatic cell reprogramming, but the underlying mechanism is largely unknown. Here, we reported that knockdown of histone deacetylase 2 (HDAC2) specifically promoted the maturation of iPS cells. Further studies showed that HDAC2 knockdown significantly increased histone acetylation, facilitated TET1 binding and DNA demethylation at the promoters of iPS cell maturation-related genes during the transition of pre-iPS cells to a fully reprogrammed state. We also found that HDAC2 competed with TET1 in the binding of the RbAp46 protein at the promoters of maturation genes and knockdown of TET1 markedly prevented the activation of these genes. Collectively, our data not only demonstrated a novel intrinsic mechanism that the HDAC2-TET1 switch critically regulates iPS cell maturation, but also revealed an underlying mechanism of the interplay between histone acetylation and DNA demethylation in gene regulation.

Ninety-Day Subchronic Oral Toxicity Study of Senecio scandens Extract in Rats.

The present study assessed the safety/toxicity of Senecio scandens, a well-known Chinese herb that is used as an anti-inflammatory, antibiosis, and antipyretic drug. A 90-d subchronic oral toxicity study of S. scandens was performed in Wistar rats. The extract of S. scandens was administered orally to male and female rats at a single dose of 225, 450, and 900 mg/kg/d. There was no obvious toxicity. Certain changes in hematology and coagulation parameters (red cell distribution width (RDW), platelet count (PLT), monocyte percentage (Mo%), activated partial thromboplastin time (APTT), prothrombin time (PT)) were observed in some administration groups. In regards to the blood biochemical parameters, the levels of creatinine (CRN), potassium, and chloride were increased in a number of the treated rats. There were no significant changes in other hematology, coagulation, or biochemical parameters in rats orally administered S. scandens. S. scandens has a slight effect on rat coagulation and metabolism systems. The herb was safe at all doses tested, but caution should be taken when administering S. scandens at higher doses.

In vivo antibacterial activity of MRX-I, a new oxazolidinone.

MRX-I is a potent oxazolidinone antibiotic against Gram-positive pathogens, including methicillin-resistant Staphylococcus aureus (MRSA), penicillin-resistant Streptococcus pneumoniae (PRSP), penicillin-intermediate S. pneumoniae (PISP), and vancomycin-resistant enterococci (VRE). In this study, the in vivo efficacy of orally administered MRX-I was evaluated using linezolid as a comparator. MRX-I showed the same or better efficacy than linezolid in both systemic and local infection models against the tested strains.

[Correlation study on Chinese medical syndrome types of chronic hepatitis B patients and HLA-DR13 gene, BCP mutation, and T-lymphocyte subsets].

OBJECTIVE: To explore the correlation between the HLA-DR13, basic core promoter (BCP), changes of T lymphocyte subset and clinical Chinese medical syndromes of chronic hepatitis B (CHB). METHODS: Totally 102 CHB patients were syndrome typed as Gan depression Pi deficiency syndrome (GDPDS), Pi-Shen yang deficiency syndrome (PSYDS), Gan-gallbladder dampness heat syndrome (GGDHS), Gan-Shen yin deficiency syndrome (GSYDS), and static blood blocking collaterals syndrome (SBBCS). Besides, 30 healthy subjects were recruited as the normal control group. The blood HBV-DNA level and HLA-DR13 gene were detected with real time fluorescent PCR. The expression of CD4+ and CD8+ in T lymphocytes was detected using flow cytometry. The mutation of serum A1762T/G1764A was detected using PCR sequencing. Hepatitis Be antigen (HBeAg) was detected with ELISA, and correlation between various Chinese medical syndrome types and objective indicators were analyzed. RESULTS: There was no statistical difference in HBV-DNA quantitative results among various syndrome types (P > 0.05). HBeAg positive rate was higher in GDPDS than in other syndrome types (P < 0.05). It was sequenced as GDPDS > GSYDS > SBBCS > GGDHS > PSYDS. Compared with the normal control group, percentages of CD3+ and CD3+ CD4+ were lower in PSYDS (P < 0.05). The ratio of CD3+ CD4+/CD3+ CD8 was lower in GGDHS and PSYDS than in the normal control group (P < 0.05). There was no statistical difference in the CD3+ CD8+ percentage among various syndrome types (P > 0.05). The quantitation of HLA-DR13 gene was lower in GDPDS and GSYDS than in the normal control group (P < 0.05). The positive rate of BCP mutation was higher in GSYDS than in other syndrome types (P < 0.05). CONCLUSION: Co-detection results of HLA-DR13 and BCP could be used as reference indices of Chinese medical syndrome typing of CHB.

Acoustic modulation and non-contact atomization of droplets based on the Fabry-Pérot resonator.

A non-contact ultrasonic atomization based on the Fabry-Pérot resonator is proposed to obtain atomized droplets with a reduced droplet diameter and concentrated droplet distributions. To better understand the mechanism inside the acoustic chamber, the acoustic-fluid interactions are numerically explored inside the Fabry-Pérot resonator to achieve the precise modulation of droplets. The influence of the acoustic chamber's geometry and the ultrasonic properties on the atomized droplet diameter and distributions is investigated, aiming to establish matching relationships between the atomized droplet diameter and the geometry of the acoustic chamber. The dynamic behaviors of droplet breakup are observed with a high-speed camera to reveal the atomization mechanism of liquid droplets in high-intensity acoustic fields. The experiments demonstrate that the proposed non-contact atomization can achieve atomized water droplets with a median diameter of ∼24 µm, providing an alternative to ultrasonic spray.

Negatively charged nanodiscs for the reduction of toxicity and enhanced efficacy of polymyxin B against Acinetobacter baumannii sepsis.

The treatment of sepsis caused by multidrug-resistant (MDR) Gram-negative bacterial infections remains challenging. With these pathogens exhibiting resistance to carbapenems and new generation cephalosporins, the traditional antibiotic polymyxin B (PMB) has reemerged as a critical treatment option. However, its severe neurotoxicity and nephrotoxicity greatly limit the clinical application. Therefore, we designed negatively charged high-density lipoprotein (HDL) mimicking nanodiscs as a PMB delivery system, which can simultaneously reduce toxicity and enhance drug efficacy. The negative charge prevented the PMB release in physiological conditions and binding to cell membranes, significantly reducing toxicity in mammalian cells and mice. Notably, nanodisc-PMB exhibits superior efficacy than free PMB in sepsis induced by carbapenem-resistant Acinetobacter baumannii (CRAB) strains. Nanodisc-PMB shows promise as a treatment for carbapenem-resistant Gram-negative bacterial sepsis, especially caused by Acinetobacter baumannii, and the nanodiscs could be repurposed for other toxic antibiotics as an innovative delivery system. STATEMENT OF SIGNIFICANCE: Multidrug-resistant Gram-negative bacteria, notably carbapenem-resistant Acinetobacter baumannii, currently pose a substantial challenge due to the scarcity of effective treatments, rendering Polymyxins a last-resort antibiotic option. However, their therapeutic application is significantly limited by severe neurotoxic and nephrotoxic side effects. Prevailing polymyxin delivery systems focus on either reducing toxicity or enhancing bioavailability yet fail to simultaneously achieve both. In this scenario, we have developed a distinctive HDL-mimicking nanodisc for polymyxin B, which not only significantly reduces toxicity but also improves efficacy against Gram-negative bacteria, especially in sepsis caused by CRAB. This research offers an innovative drug delivery system for polymyxin B. Such advancement could notably improve the therapeutic landscape and make a significant contribution to the arsenal against these notorious pathogens.

The interaction of endorepellin and neurexin triggers neuroepithelial autophagy and maintains neural tube development.

Heparan sulfate proteoglycan 2 (HSPG2) gene encodes the matrix protein Perlecan, and genetic inactivation of this gene creates mice that are embryonic lethal with severe neural tube defects (NTDs). We discovered rare genetic variants of HSPG2 in 10% cases compared to only 4% in controls among a cohort of 369 NTDs. Endorepellin, a peptide cleaved from the domain V of Perlecan, is known to promote angiogenesis and autophagy in endothelial cells. The roles of enderepellin in neurodevelopment remain unclear so far. Our study revealed that endorepellin can migrate to the neuroepithelial cells and then be recognized and bind with the neuroepithelia receptor neurexin in vivo. Through the endocytic pathway, the interaction of endorepellin and neurexin physiologically triggers autophagy and appropriately modulates the differentiation of neural stem cells into neurons as a blocker, which is necessary for normal neural tube closure. We created knock-in (KI) mouse models with human-derived HSPG2 variants, using sperm-like stem cells that had been genetically edited by CRISPR/Cas9. We realized that any HSPG2 variants that affected the function of endorepellin were considered pathogenic causal variants for human NTDs given that the severe NTD phenotypes exhibited by these KI embryos occurred in a significantly higher response frequency compared to wildtype embryos. Our study provides a paradigm for effectively confirming pathogenic mutations in other genetic diseases. Furthermore, we demonstrated that using autophagy inhibitors at a cellular level can repress neuronal differentiation. Therefore, autophagy agonists may prevent NTDs resulting from failed autophagy maintenance and neuronal over-differentiation caused by deleterious endorepellin variants.