The choroid plexus links innate immunity to CSF dysregulation in hydrocephalus.

The choroid plexus (ChP) is the blood-cerebrospinal fluid (CSF) barrier and the primary source of CSF. Acquired hydrocephalus, caused by brain infection or hemorrhage, lacks drug treatments due to obscure pathobiology. Our integrated, multi-omic investigation of post-infectious hydrocephalus (PIH) and post-hemorrhagic hydrocephalus (PHH) models revealed that lipopolysaccharide and blood breakdown products trigger highly similar TLR4-dependent immune responses at the ChP-CSF interface. The resulting CSF "cytokine storm", elicited from peripherally derived and border-associated ChP macrophages, causes increased CSF production from ChP epithelial cells via phospho-activation of the TNF-receptor-associated kinase SPAK, which serves as a regulatory scaffold of a multi-ion transporter protein complex. Genetic or pharmacological immunomodulation prevents PIH and PHH by antagonizing SPAK-dependent CSF hypersecretion. These results reveal the ChP as a dynamic, cellularly heterogeneous tissue with highly regulated immune-secretory capacity, expand our understanding of ChP immune-epithelial cell cross talk, and reframe PIH and PHH as related neuroimmune disorders vulnerable to small molecule pharmacotherapy.

The mechano-chemical circuit drives skin organoid self-organization.

Stem cells in organoids self-organize into tissue patterns with unknown mechanisms. Here, we use skin organoids to analyze this process. Cell behavior videos show that the morphological transformation from multiple spheroidal units with morphogenesis competence (CMU) to planar skin is characterized by two abrupt cell motility-increasing events before calming down. The self-organizing processes are controlled by a morphogenetic module composed of molecular sensors, modulators, and executers. Increasing dermal stiffness provides the initial driving force (driver) which activates Yap1 (sensor) in epidermal cysts. Notch signaling (modulator 1) in epidermal cyst tunes the threshold of Yap1 activation. Activated Yap1 induces Wnts and MMPs (epidermal executers) in basal cells to facilitate cellular flows, allowing epidermal cells to protrude out from the CMU. Dermal cell-expressed Rock (dermal executer) generates a stiff force bridge between two CMU and accelerates tissue mixing via activating Laminin and ß1-integrin. Thus, this self-organizing coalescence process is controlled by a mechano-chemical circuit. Beyond skin, self-organization in organoids may use similar mechano-chemical circuit structures.

High-power 100 W Kerr-lens mode-locked ring-cavity femtosecond Yb:YAG thin-disk oscillator.

High-power femtosecond pulses delivered at a high-repetition rate will aid machining throughput and improve signal-to-noise ratios for sensitive measurements. Here we demonstrate a Kerr-lens mode-locked femtosecond Yb:YAG ring-cavity thin-disk oscillator with a multi-pass scheme for the laser beam. With four passes through the thin disk, 175-fs pulses were delivered from the oscillator at an average power of 71.5 W and a repetition rate of 65.3 MHz. The corresponding intra-cavity peak power of 110 MW is ample for intra-cavity nonlinear conversion into more exotic wavelength ranges. With six passes, the average output power reached 101.3 W. To the best of our knowledge, this is the highest average output power of any mode-locked ring laser. These results confirm the viability of using multi-pass configuration on a thin-disk ring oscillator for high-throughput femtosecond applications.

Time trends and regional variation in utilization of antidiabetic medicines in China, 2015-2022.

AIM: To assess the use of non-insulin antidiabetic medicines in China. MATERIALS AND METHODS: We analysed the national procurement data for 29 non-insulin antidiabetic medicines from nine subgroups in China from 2015 to 2022. We estimated the number of defined daily doses (DDDs) procured per year in seven regions of China for nine subgroups and adjusted the data by the number of patients with diabetes. For each subgroup, the regional ratio was calculated by comparing the procurement per patient in each region with the procurement nationwide. The regional disparity was the difference between the highest and lowest regional ratios. We compared the medication patterns across regions. RESULTS: Nationally, between 2015 and 2022, the number of DDDs per patient increased from 14.45 to 47.37. The two most commonly used categories were sulphonylurea and biguanides, which increased from 7.04 to 15.39 (119%) and 3.28 to 11.11 (239%) DDDs per patient, respectively. The procurement of new drugs (dipeptidyl peptidase-4 inhibitors, sodium-glucose cotransporter type 2 inhibitors and glucagon-like peptide-1 receptor agonists) increased quickly and had >5000% relative changes. Particularly for sodium-glucose cotransporter type 2 inhibitors, it increased from 0.08 to 5.03 DDDs (6662%). The southwest region had the highest relative change (319%), while the southern region had the lowest (118%). Biguanide and thiazolidinediones had the lowest (1.19) and highest level (2.21) of regional disparity in 2022, respectively. CONCLUSION: The procurement of non-insulin antidiabetic medicines in China has increased a lot from 2015 to 2022. In terms of DDDs per patient, sulphonylurea ranked first, followed by metformin. The procurement of new drugs increased greatly. A large regional disparity existed in medicine usage and patterns.

Hereditary causes of hypertension due to increased sodium transport.

PURPOSE OF REVIEW: Hypertension, commonly known as high blood pressure, is a widespread health condition affecting a large number of individuals across the globe. Although lifestyle choices and environmental factors are known to have a significant impact on its development, there is growing recognition of the influence of genetic factors in the pathogenesis of hypertension. This review specifically focuses on the hereditary causes of hypertension that are associated with increased sodium transport through the thiazide-sensitive NaCl cotransporter (NCC) or amiloride-sensitive epithelial sodium channel (ENaC), crucial mechanisms involved in regulating blood pressure in the kidneys. By examining genetic mutations and signaling molecules linked to the dysregulation of sodium transport, this review aims to deepen our understanding of the hereditary causes of hypertension and shed light on potential therapeutic targets. RECENT FINDINGS: Liddle syndrome (LS) is a genetic disorder that typically manifests early in life and is characterized by hypertension, hypokalemic metabolic alkalosis, hyporeninemia, and suppressed aldosterone secretion. This condition is primarily caused by gain-of-function mutations in ENaC. In contrast, Pseudohypoaldosteronism type II (PHAII) is marked by hyperkalemia and hypertension, alongside other clinical features such as hyperchloremia, metabolic acidosis, and suppressed plasma renin levels. PHAII results from overactivations of NCC, brought about by gain-of-function mutations in its upstream signaling molecules, including WNK1 (with no lysine (K) 1), WNK4, Kelch-like 3 (KLHL3), and cullin3 (CUL3). SUMMARY: NCC and ENaC are integral components, and their malfunctions lead to disorders like LS and PHAII, hereditary causes of hypertension. Current treatments for LS involve ENaC blockers (e.g., triamterene and amiloride) in conjunction with low-sodium diets, effectively normalizing blood pressure and potassium levels. In PHAII, thiazide diuretics, which inhibit NCC, are the mainstay treatment, albeit with some limitations and potential side effects. Ongoing research in developing alternative treatments, including small molecules targeting key regulators, holds promise for more effective and tailored hypertension solutions.

N6-Methyladenosine modified circ-NAB1 modulates cell cycle and epithelial-mesenchymal transition via CDKN3 in endometrial cancer.

Endometrial cancer (EC) is a common malignant tumor in the female reproductive system. Circular RNAs (circRNAs) and N6-methyladenosine (m6A) modification are widely involved in cancer progression. Nevertheless, the cross-talk between circ-NAB1 and m6A as well as the biological functions of circ-NAB1 in EC remain unclear. Circ-NAB1 was observed to be upregulated in EC tissues and cells by RT-qPCR. MeRIP and RNA pull-down assays were utilized for detecting the m6A modification of circ-NAB1. The interaction between circ-NAB1 and RNAs was also detected. Colony formation, transwell, flow cytometry, and western blot were utilized for measuring EC cell behaviors. Mechanically, we proved the m6A demethylase alkylation repair homolog protein 5 (ALKBH5) can mediate circ-NAB1 expression through an m6A-YTHDF2-dependent manner. Circ-NAB1 overexpression can promote cell proliferation, migration, invasion, epithelial-mesenchymal transition (EMT) process, and cell cycle through functional assays. Circ-NAB1 knockdown exerts the opposite function on EC cells. Furthermore, we proved that circ-NAB1 can sponge miR-876-3p to upregulate the target gene cyclin-dependent kinase inhibitor 3 (CDKN3) in EC cells. CDKN3 overexpression can reverse the impacts of circ-NAB1 depletion on EC cell behaviors. Collectively, we proved that ALKBH5-mediated m6A modification of circ-NAB1 promoted EMT process and cell cycle in EC via targeting the miR-876-3p/CDKN3 axis.

Tanshinone IIA regulates CCl4 induced liver fibrosis in C57BL/6J mice via the PI3K/Akt and Nrf2/HO-1 signaling pathways.

Chronic liver diseases caused by various factors may develop into liver fibrosis (LF). Early stage of LF could be reversible. Tanshinone IIA (Tan IIA), an extract from Salvia miltiorrhiza, has been reported to be hepatoprotective. However, the potential targets and mechanism of Tan IIA in the treatment of LF are still unclear. Our study aims at the anti-LF mechanism of Tan IIA through network pharmacological analysis combined with LF-related experiments. Serum biochemical indicators and histopathological examination showed that Tan IIA could ameliorate the process of LF in the CCl4 -induced mouse model. Western blot and immunohistochemical assays showed that Tan IIA decreased the expression of Kirsten rat sarcoma viral oncogene homolog (KRAS), phosphatidylinositide 3-kinases/protein kinase B (PI3K/Akt), and nuclear factor erythroid 2-related factor/heme oxygenase-1 (Nrf2/HO-1). Compared with the model group, the Tan IIA groups increased the decreased superoxide dismutase activity and glutathione content, while decreasing the increased malondialdehyde content. These results indicate that Tan IIA may play an antioxidant role by inhibiting the expression of KRAS, PI3K/Akt, and Nrf2/HO-1 to ameliorate the progression of LF, which to some extent explains the pharmacological mechanism of Tan IIA in LF. In conclusion, our study demonstrates that Tan IIA could regulate LF via PI3K/Akt and Nrf2/HO-1 signaling pathways. It may be an effective therapeutic compound for the treatment of LF.

Neuronal K+-Cl- cotransporter KCC2 as a promising drug target for epilepsy treatment.

Epilepsy is a prevalent neurological disorder characterized by unprovoked seizures. γ-Aminobutyric acid (GABA) serves as the primary fast inhibitory neurotransmitter in the brain, and GABA binding to the GABAA receptor (GABAAR) regulates Cl- and bicarbonate (HCO3-) influx or efflux through the channel pore, leading to GABAergic inhibition or excitation, respectively. The neuron-specific K+-Cl- cotransporter 2 (KCC2) is essential for maintaining a low intracellular Cl- concentration, ensuring GABAAR-mediated inhibition. Impaired KCC2 function results in GABAergic excitation associated with epileptic activity. Loss-of-function mutations and altered expression of KCC2 lead to elevated [Cl-]i and compromised synaptic inhibition, contributing to epilepsy pathogenesis in human patients. KCC2 antagonism studies demonstrate the necessity of limiting neuronal hyperexcitability within the brain, as reduced KCC2 functioning leads to seizure activity. Strategies focusing on direct (enhancing KCC2 activation) and indirect KCC2 modulation (altering KCC2 phosphorylation and transcription) have proven effective in attenuating seizure severity and exhibiting anti-convulsant properties. These findings highlight KCC2 as a promising therapeutic target for treating epilepsy. Recent advances in understanding KCC2 regulatory mechanisms, particularly via signaling pathways such as WNK, PKC, BDNF, and its receptor TrkB, have led to the discovery of novel small molecules that modulate KCC2. Inhibiting WNK kinase or utilizing newly discovered KCC2 agonists has demonstrated KCC2 activation and seizure attenuation in animal models. This review discusses the role of KCC2 in epilepsy and evaluates its potential as a drug target for epilepsy treatment by exploring various strategies to regulate KCC2 activity.

Lineage-specific insertions in T-box riboswitches modulate antibiotic binding and action.

T-box riboswitches (T-boxes) are essential RNA regulatory elements with a remarkable structural diversity, especially among bacterial pathogens. In staphylococci, all glyS T-boxes synchronize glycine supply during synthesis of nascent polypeptides and cell wall formation and are characterized by a conserved and unique insertion in their antiterminator/terminator domain, termed stem Sa. Interestingly, in Staphylococcus aureus the stem Sa can accommodate binding of specific antibiotics, which in turn induce robust and diverse effects on T-box-mediated transcription. In the present study, domain swap mutagenesis and probing analysis were performed to decipher the role of stem Sa. Deletion of stem Sa significantly reduces both the S. aureus glyS T-box-mediated transcription readthrough levels and the ability to discriminate among tRNAGly isoacceptors, both in vitro and in vivo. Moreover, the deletion inverted the previously reported stimulatory effects of specific antibiotics. Interestingly, stem Sa insertion in the terminator/antiterminator domain of Geobacillus kaustophilus glyS T-box, which lacks this domain, resulted in elevated transcription in the presence of tigecycline and facilitated discrimination among proteinogenic and nonproteinogenic tRNAGly isoacceptors. Overall, stem Sa represents a lineage-specific structural feature required for efficient staphylococcal glyS T-box-mediated transcription and it could serve as a species-selective druggable target through its ability to modulate antibiotic binding.

Anticancer medicines in Pakistan: An analysis of essential medicines lists.

OBJECTIVE: The lack of anticancer drugs for curative and supportive purposes is the critical reason for the low survival rate in low-and-middle-income countries. This study aims to analyze whether the National Essential Medicines List (NEML) and Registered Essential Medicines List (REML) are in concordance with the World Health Organization (WHO) Essential Medicines List (EML) and whether the formularies prevalent in the country are parallel to each other and to the NEML. METHOD: An observational study design was used in which antineoplastic drugs from the 2021 NEML and REML were compared with 2021 WHO EML to evaluate their availability in Pakistan. Market access was determined. Moreover, the formularies of six different hospital types were compared with each other and with the NEML, and REML to estimate the availability within hospitals. RESULTS: There were 66 anticancer drugs in 2021 WHO EML and all were found in Pakistan's 2021 NEML but only 48 drugs (73%) were found in the REML. Hydroxycarbamide and dasatinib were two registered drugs absent in all hospitals' formularies. The market access for anticancer medicines was 73% (48 of 66). Semigovernment hospital (86%) has the highest availability, followed by the government hospital (80%). All the hospitals have unregistered drugs including bortezomib, lenalidomide, and mesna. CONCLUSION: Pakistan's NEML adopts WHO EML abruptly but all medicines are not registered. The hospitals are trying their best to increase availability but optimum drug regulations to revise NEML based on the country's requirements and emphasizing registration of anticancer medicines are needed to improve the country's availability of antineoplastic agents.

Intelligent Path Planning with an Improved Sparrow Search Algorithm for Workshop UAV Inspection.

Intelligent workshop UAV inspection path planning is a typical indoor UAV path planning technology. The UAV can conduct intelligent inspection on each work area of the workshop to solve or provide timely feedback on problems in the work area. The sparrow search algorithm (SSA), as a novel swarm intelligence optimization algorithm, has been proven to have good optimization performance. However, the reduction in the SSA's search capability in the middle or late stage of iterations reduces population diversity, leading to shortcomings of the algorithm, including low convergence speed, low solution accuracy and an increased risk of falling into local optima. To overcome these difficulties, an improved sparrow search algorithm (namely the chaotic mapping-firefly sparrow search algorithm (CFSSA)) is proposed by integrating chaotic cube mapping initialization, firefly algorithm disturbance search and tent chaos mapping perturbation search. First, chaotic cube mapping was used to initialize the population to improve the distribution quality and diversity of the population. Then, after the sparrow search, the firefly algorithm disturbance and tent chaos mapping perturbation were employed to update the positions of all individuals in the population to enable a full search of the algorithm in the solution space. This technique can effectively avoid falling into local optima and improve the convergence speed and solution accuracy. The simulation results showed that, compared with the traditional intelligent bionic algorithms, the optimized algorithm provided a greatly improved convergence capability. The feasibility of the proposed algorithm was validated with a final simulation test. Compared with other SSA optimization algorithms, the results show that the CFSSA has the best efficiency. In an inspection path planning problem, the CFSSA has its advantages and applicability and is an applicable algorithm compared to SSA optimization algorithms.

Regulation of Neuronal Chloride Homeostasis by Pro- and Mature Brain-Derived Neurotrophic Factor (BDNF) via KCC2 Cation-Chloride Cotransporters in Rat Cortical Neurons.

The strength of inhibitory neurotransmission depends on intracellular neuronal chloride concentration, primarily regulated by the activity of cation-chloride cotransporters NKCC1 (Sodium-Potassium-Chloride Cotransporter 1) and KCC2 (Potassium-Chloride Cotransporter 2). Brain-derived neurotrophic factor (BDNF) influences the functioning of these co-transporters. BDNF is synthesized from precursor proteins (proBDNF), which undergo proteolytic cleavage to yield mature BDNF (mBDNF). While previous studies have indicated the involvement of BDNF signaling in the activity of KCC2, its specific mechanisms are unclear. We investigated the interplay between both forms of BDNF and chloride homeostasis in rat hippocampal neurons and in utero electroporated cortices of rat pups, spanning the behavioral, cellular, and molecular levels. We found that both pro- and mBDNF play a comparable role in immature neurons by inhibiting the capacity of neurons to extrude chloride. Additionally, proBDNF increases the endocytosis of KCC2 while maintaining a depolarizing shift of EGABA in maturing neurons. Behaviorally, proBDNF-electroporated rat pups in the somatosensory cortex exhibit sensory deficits, delayed huddling, and cliff avoidance. These findings emphasize the role of BDNF signaling in regulating chloride transport through the modulation of KCC2. In summary, this study provides valuable insights into the intricate interplay between BDNF, chloride homeostasis, and inhibitory synaptic transmission, shedding light on the underlying cellular mechanisms involved.

Acetate Alleviates Gut Microbiota Depletion-Induced Retardation of Skeletal Muscle Growth and Development in Young Mice.

The normal growth and development of skeletal muscle is essential for the health of the body. The regulation of skeletal muscle by intestinal microorganisms and their metabolites has been continuously demonstrated. Acetate is the predominant short-chain fatty acids synthesized by gut microbiota through the fermentation of dietary fiber; however, the underlying molecular mechanisms governing the interaction between acetate and skeletal muscle during the rapid growth stage remains to be further elucidated. Herein, specific pathogen-free (SPF) mice, germ-free (GF) mice, and germ-free mice supplemented with sodium acetate (GS) were used to evaluate the effects of acetate on the skeletal muscle growth and development of young mice with gut microbiota deficiency. We found that the concentration of serum acetate, body mass gain, succinate dehydrogenase activity, and expression of the myogenesis maker gene of skeletal muscle in the GS group were higher than those in the GF group, following sodium acetate supplementation. Furthermore, the transcriptome analysis revealed that acetate activated the biological processes that regulate skeletal muscle growth and development in the GF group, which are otherwise inhibited due to a gut microbiota deficiency. The in vitro experiment showed that acetate up-regulated Gm16062 to promote skeletal muscle cell differentiation. Overall, our findings proved that acetate promotes skeletal muscle growth and development in young mice via increasing Gm16062 expression.

Dexmedetomidine adjunct to ropivacaine for ultrasound-guided transversus abdominis plane block for open inguinal hernia repair in the older adults: A randomised clinical trial.

OBJECTIVE: The aim of this study was to evaluate the effect of adding dexmedetomidine to ropivacaine on pain relief and quality of recovery in older patients undergoing open inguinal hernia repair surgeries. METHODS: This was a prospective and randomised clinical trial of 102 patients aged over 65 years who received an ultrasound-guided transversus open mesh herniorrhaphy abdominis plane (TAP) block with either 0.375% ropivacaine 20 ml (Group R, n = 47) or 0.375% ropivacaine combined with 1 µg/kg dexmedetomidine 20 ml (Group RD, n = 45) in the pre-anaesthesia care unit before elective open inguinal hernia surgeries. The primary outcome measure was Visual Analogue Scale (VAS) pain scores at rest and on movement at 2, 4, 8, 12 and 24 h and at 1 and 3 months' postoperatively. The secondary outcome measures were the incidence of post-operative delirium (POD), nausea and vomiting and the occurrence of side effects or complications on post-operative day 1. RESULTS: Group RD had lower VAS scores at rest and on movement at 8 and 12 h postoperatively and a lower incidence of POD on the post-operative day 1 than Group R. Transient bradycardia was more frequent in Group RD than in Group R, and side effects or post-operative complications were reported in either group. CONCLUSION: The addition of dexmedetomidine to ropivacaine in a TAP block enhances postoperative analgesia during hospitalisation and improves the quality of recovery without affecting chronic pain in older patients undergoing open inguinal hernia repair surgery.

Turning on Singlet Oxygen Generation by Outer-Sphere Microenvironment Modulation in Porphyrinic Covalent Organic Frameworks for Photocatalytic Oxidation.

Singlet oxygen (1 O2 ) is ubiquitously involved in various photocatalytic oxidation reactions; however, efficient and selective production of 1 O2 is still challenging. Herein, we reported the synthesis of nickel porphyrin-based covalent organic frameworks (COFs) incorporating functional groups with different electron-donating/-withdrawing features on their pore walls. These functional groups established a dedicated outer-sphere microenvironment surrounding the Ni catalytic center that tunes the activity of the COFs for 1 O2 -mediated thioether oxidation. With the increase of the electron-donating ability of functional groups, the modulated outer-sphere microenvironment turns on the catalytic activity from a yield of nearly zero by the cyano group functionalized COF to an excellent yield of 98 % by the methoxy group functionalized one. Electronic property investigation and density-functional theory (DFT) calculations suggested that the distinct excitonic behaviors attributed to the diverse band energy levels and orbital compositions are responsible for the different activities. This study represents the first regulation of generating reactive oxygen species (ROS) based on the strategy of outer-sphere microenvironment modulation in COFs.

High-Level Secretion of Pregnancy Zone Protein Is a Novel Biomarker of DNA Damage-Induced Senescence and Promotes Spontaneous Senescence.

Identification of unique and specific biomarkers to better detect and quantify senescent cells remains challenging. By a global proteomic profiling of senescent human skin BJ fibroblasts induced by ionizing radiation (IR), the cellular level of pregnancy zone protein (PZP), a presumable pan-protease inhibitor never been linked to cellular senescence before, was found to be decreased by more than 10-fold, while the level of PZP in the conditioned medium was increased concomitantly. This observation was confirmed in a variety of senescent cells induced by IR or DNA-damaging drugs, indicating that high-level secretion of PZP is a novel senescence-associated secretory phenotype. RT-PCR examination verified that the transcription of the PZP gene is enhanced in various cells at senescence or upregulated following DNA damage treatment in a p53-independent manner. Moreover, pretreatment with late pregnancy serum containing a high level of PZP led to inhibition of doxorubicin-induced senescence in A549 cells, and depletion of PZP in the pregnancy serum could enhance such inhibition. Finally, the addition of immuno-precipitated PZP complexes into tissue culture attenuated the growth of A549 cells and promoted the spontaneous senescence. Therefore, we revealed that high-level secretion of PZP is a novel and unique feature associated with DNA damage-induced senescence, and secreted PZP is a positive regulator of cellular senescence, particularly during the late stage of gestation.

LARO: Learned acquisition and reconstruction optimization to accelerate quantitative susceptibility mapping.

Quantitative susceptibility mapping (QSM) involves acquisition and reconstruction of a series of images at multi-echo time points to estimate tissue field, which prolongs scan time and requires specific reconstruction technique. In this paper, we present our new framework, called Learned Acquisition and Reconstruction Optimization (LARO), which aims to accelerate the multi-echo gradient echo (mGRE) pulse sequence for QSM. Our approach involves optimizing a Cartesian multi-echo k-space sampling pattern with a deep reconstruction network. Next, this optimized sampling pattern was implemented in an mGRE sequence using Cartesian fan-beam k-space segmenting and ordering for prospective scans. Furthermore, we propose to insert a recurrent temporal feature fusion module into the reconstruction network to capture signal redundancies along echo time. Our ablation studies show that both the optimized sampling pattern and proposed reconstruction strategy help improve the quality of the multi-echo image reconstructions. Generalization experiments show that LARO is robust on the test data with new pathologies and different sequence parameters. Our code is available at https://github.com/Jinwei1209/LARO-QSM.git.

Combinatorial Drug Screening Based on Massive 3D Tumor Cultures Using Micropatterned Array Chips.

The establishment and application of a generalizable three-dimensional (3D) tumor device for high-throughput screening plays an important role in drug discovery and cancer therapeutics. In this study, we introduce a facile microplatform for considerable 3D tumor generation and combinatorial drug screening evaluation. High fidelity of chip fabrication was achieved depending on the simple and well-improved microcontact printing. We demonstrated the high stability and repeatability of the established tumor-on-a-chip system for controllable and massive production of 3D tumors with high size uniformity. Importantly, we accomplished the screening-like chemotherapy investigation involving individual and combinatorial drugs and validated the high accessibility and applicability of the system in 3D tumor-based manipulation and analysis on a large scale. This achievement in tumor-on-a-chip has potential applications in plenty of biomedical fields such as tumor biology, pharmacology, and tissue microengineering. It offers an insight into the development of the popularized microplatform with easy-to-fabricate and easy-to-operate properties for cancer exploration and therapy.

KIR2DL4/HLA-G polymorphisms were associated with HCV infection susceptibility among Chinese high-risk population.

Killer-cell immunoglobulin-like receptors 2DL4 (KIR2DL4) and the human leukocyte antigen class I-G (HLA-G) display vital parts in immune responses against hepatitis C virus (HCV) infection. We select four potentially functional single nucleotide polymorphisms (SNPs) of KIR/HLA to explore the associations between KIR2DL4/HLA-G genetic variants and HCV infection results. In the present case-control study, a total of 2225 HCV-infected high-risk subjects, including 1778 paid blood donors (PBD) and 447 drug users were consecutively recruited before treatment from 2011 to 2018. KIR2DL4-rs660773, KIR2DL4-rs660437, HLA-G-rs9380142, and HLA-G-rs1707 SNPs were sorted as genotypes in the subdivided groups, involving 1095 uninfected controls subjects, 432 spontaneous HCV clearance subjects and 698 HCV persistent infection subjects. After genotyping experiments using the TaqMan-MGB assay, modified logistic regression was used to calculate the correlation among the SNPs and HCV infection. The SNPs were functionally annotated using bioinformatics analysis. Following adjusting by age, sex, alanine aminotransferase, aspartate aminotransferase, IFNL3-rs12979860, IFNL3-rs8099917, and the infection route, the logistic regression analysis discovered that KIR2DL4-rs660773 and HLA-G-rs9380142 were correlated with vulnerability to HCV infection (all p < 0.05). In a locus-dosage way, compared with subjects carrying the rs9380142-AA or rs660773-AA genotypes, subjects with rs9380142-AG or rs660773-AG/GG (all p < 0.05) were more vulnerable to HCV infection; the overall impact of their risk genotypes (rs9380142-AGrs660773-AG/GG) was correlated with an elevated incidence of HCV infection (ptrend < 0.001). In the Haplotype analysis, patients with haplotype AG were more likely to contract HCV compared to those with the highest common AA haplotype (p = 0.002) were higher in susceptibility to infect HCV. The SNPinfo web server estimated that rs660773 is a transcription factor binding site, whereas rs9380142 is a potential microRNA-binding site. In two Chinese high-risk population (PBD and drug uesrs), KIR2DL4 rs660773-G and HLA-G rs9380142-G alleles polymorphisms are related to HCV susceptibility. KIR2DL4/HLA-G pathway genes might affect the innate immune responses by regulating KIR2DL4/HLA-G transcription and translation play a potential role in HCV infection.

S9.6 Antibody-Enzyme Conjugates for the Detection of DNA-RNA Hybrids.

Diagnosis of infectious agents is increasingly done by the detection of unique nucleic acid sequences, typically using methods such as PCR that specifically amplify these sequences. A largely neglected alternative approach is to use antibodies that recognize nucleic acids. The unique monoclonal antibody S9.6 recognizes DNA-RNA hybrids in a largely sequence-independent manner. S9.6 has been used in several cases for the analysis of nucleic acids. Extending our recent determination of the structure of S9.6 Fab bound to a DNA-RNA hybrid, we have developed reagents and methods for the sensitive detection of specific DNA and RNA sequences. To facilitate the use in diagnostics, we conjugated the S9.6 Fab to the highly active and well-characterized reporter enzyme human-secreted embryonic alkaline phosphatase (SEAP). Two approaches were utilized for conjugation. The first used sortase A (SrtA), which generates a covalent peptide bond between short amino acid sequences added to recombinantly produced S9.6 Fab and SEAP. The second approach was to genetically fuse the S9.6 Fab and SEAP so that the two are produced as a single molecule. Using these two antibody-SEAP proteins, we developed a simplified ELISA format for the identification of synthetic DNA-RNA hybrids, which can be optimized for detecting nucleic acids of pathogens, as well as for other applications. We successfully used this immunosorbent assay, HC-S, to identify DNA-RNA hybrids in solution with high specificity and sensitivity.