Dynamic budget analysis of multiple parameters in a lithography system based on the superposition of light intensity fluctuations.

Lithography is one of the most critical processes in the manufacturing of micro- and nano-devices. As device critical dimensions continue to shrink, variations in system parameters during the lithography process often result in heavy deviations from the intended targets, making control of these parameters crucial to ensure that lithography results meet process requirements. Gaining a thorough comprehension of how various parameters interact and contribute to lithography errors is significant, and it is equally important to offer precise suggestions for managing these parameters in extreme ultraviolet lithography (EUVL) scanners. This paper analyzes the key physical factors in the light source, illumination system and projection system of EUVL scanners and proposes what we believe to be a new methodology of budget analysis utilizing the superposition of light intensity fluctuations. Then the corresponding characteristics of light intensity fluctuations are analyzed when these parameters have fluctuated through theoretical formula derivation. A mapping model was established between parameter fluctuations and imaging outcomes through the distribution of light intensity. The yield requirements for critical dimension and pattern shift in EUVL are used to determine the exact budget range for each parameter in the proposed methodology. By controlling the parameters according to the budget analysis method proposed in this paper, the deviation between the experimental results from the yield requirements is no more than 0.1% in average. This approach allows for dynamic updating of the control range of relevant parameters based on their distinct characteristics to accommodate the unique fingerprints of various EUVL scanners. Furthermore, based on this adaptive budget range of multiple parameters, it can offer distinct direction for the development of lithography equipment or serve as a clear indication for parameter monitoring.

At-Home Saliva Sampling in Healthy Adults Using CandyCollect, a Lollipop-Inspired Device.

Respiratory infections are common in children, and there is a need for user-friendly collection methods. Here, we performed the first human subjects study using the CandyCollect device, a lollipop-inspired saliva collection device .We showed that the CandyCollect device can be used to collect salivary bacteria from healthy adults using Streptococcus mutans and Staphylococcus aureus as proof-of-concept commensal bacteria. We enrolled healthy adults in a nationwide (USA) remote study in which participants were sent study packages containing CandyCollect devices and traditional commercially available oral swabs and spit tubes. Participants sampled themselves at home, completed usability and user preference surveys, and mailed the samples back to our laboratory for analysis by qPCR. Our results showed that for participants in which a given bacterium (S. mutans or S. aureus) was detected in one or both of the commercially available methods (oral swab and/or spit tubes), CandyCollect devices had a 100% concordance with the positive result (n = 14 participants). Furthermore, the CandyCollect device was ranked the highest preference sampling method among the three sampling methods by 26 participants surveyed (combining survey results across two enrollment groups). We also showed that the CandyCollect device has a shelf life of up to 1 year at room temperature, a storage period that is convenient for clinics or patients to keep the CandyCollect device and use it any time. Taken together, we have demonstrated that the CandyCollect is a user-friendly saliva collection tool that has the potential to be incorporated into diagnostic assays in clinic visits and telemedicine.

Complete Reductive Dechlorination of 4-Hydroxy-chlorothalonil by Dehalogenimonas Populations.

Chlorothalonil (2,4,5,6-tetrachloroisophthalonitrile, TePN) is one of the most widely used fungicides all over the world. Its major environmental transformation product 4-hydroxy-chlorothalonil (4-hydroxy-2,5,6-trichloroisophthalonitrile, 4-OH-TPN) is more persistent, mobile, and toxic and is frequently detected at a higher concentration in various habitats compared to its parent compound TePN. Further microbial transformation of 4-OH-TPN has never been reported. In this study, we demonstrated that 4-OH-TPN underwent complete microbial reductive dehalogenation to 4-hydroxy-isophthalonitrile via 4-hydroxy-dichloroisophthalonitrile and 4-hydroxy-monochloroisophthalonitrile. 16S rRNA gene amplicon sequencing demonstrated that Dehalogenimonas species was enriched from 6% to 17-22% after reductive dechlorination of 77.24 µmol of 4-OH-TPN. Meanwhile, Dehalogenimonas copies increased by one order of magnitude and obtained a yield of 1.78 ± 1.47 × 108 cells per µmol Cl- released (N = 6), indicating that 4-OH-TPN served as the terminal electron acceptor for organohalide respiration of Dehalogenimonas species. A draft genome of Dehalogenimonas species was assembled through metagenomic sequencing, which harbors 30 putative reductive dehalogenase genes. Syntrophobacter, Acetobacterium, and Methanosarcina spp. were found to be the major non-dechlorinating populations in the microbial community, who might play important roles in the reductive dechlorination of 4-OH-TPN by the Dehalogenimonas species. This study first reports that Dehalogenimonas sp. can also respire on the seemingly dead-end product of TePN, paving the way to complete biotransformation of the widely present TePN and broadening the substrate spectrum of Dehalogenimonas sp. to polychlorinated hydroxy-benzonitrile.

The classification of coronal deformity based on preoperative global coronal malalignment for adult spinal deformity is questionable.

STUDY DESIGN: Retrospective case-control radiographic study. OBJECTIVE: To identify main effects of preoperative pattern and global coronal malalignment (GCM) on postoperative coronal imbalance in degenerative lumbar scoliosis (DLS) patients and evaluate the rationality of the classification of coronal deformity based on preoperative GCM. A classification of coronal deformity based on preoperative GCM (20 mm set as the threshold of coronal imbalance) has been proposed recently, but whether it is practical is unclear. METHODS: One hundred twelve DLS patients treated with posterior instrumented fusion were reviewed. Coronal measurements included GCM and major Cobb angle. Based on relationship between C7 PL and major curve, preoperative patterns were classified into: Pattern 1(concave pattern), C7 PL shifted to the concave side of major curve; Pattern 2(convex pattern), C7 PL shifted to the convex side of major curve. Patients were separated into 4 groups (3 types): Type 0-1: GCM < 20 mm plus Pattern 1; Type 0-2: GCM < 20 mm plus Pattern 2; Type 1: GCM > 20 mm plus Pattern 1; Type 2: GCM > 20 mm plus Pattern 2. After comparison within patterns or among 4 groups, further factorial analysis was performed. RESULTS: Significant differences regarding postoperative GCM or coronal imbalance/balance ratio existed among 4 groups (F = 6.219, p = 0.001; x2 = 22.506, p < 0.001, respectively), despite no significant difference in intra-pattern 1(concave pattern) or intra-pattern 2(convex pattern) groups. Two-way analysis of variance showed preoperative pattern exhibited significant effect on postoperative GCM or imbalance/balance ratio (F(1,108) = 14.286, p < 0.001; F(1,108) = 30.514, p < 0.001, respectively) while neither preoperative GCM alone nor interaction of preoperative GCM with pattern did. CONCLUSION: In DLS patients, it's the preoperative pattern other than GCM that had main effects on postoperative coronal imbalance. Classification of coronal deformity based on preoperative GCM is questionable.

Rhizobium flavescens sp. nov., Isolated from a Chlorothalonil-Contaminated Soil.

A Gram-negative, facultative anaerobic, non-lagellated and rod-shaped bacterium FML-4T was isolated from a chlorothalonil-contaminated soil in Nanjing, China. Phylogenetic analyses of 16S rRNA genes revealed that the strain FML-4T shared the highest sequence similarity of 97.1% with Ciceribacter thiooxidans KCTC 52231T, followed by Rhizobium rosettiformans CCM 7583T (97.0%) and R. daejeonense KCTC 12121T (96.8%). Although the sequence similarities of the housekeeping genes thrC, rceA, glnII, and atpD between strain FML-4T and C. thiooxidans KCTC 52231T were 83.8%, 88.7%, 86.2%, and 92.0%, respectively, strain FML-4T formed a monophyletic clade in the cluster of Rhizobium species. Importantly, the feature gene of the genus Rhizobium, nifH gene (encoding the dinitrogenase reductase), was detected in strain FML-4T but not in C. thiooxidans KCTC 52231T. In addition, strain FML-4T contained the summed feature 8 (C18:1ω7c and/or C18:1ω6c), C19:0 cyclo ω8c and C16:0 as the major fatty acids. Genome sequencing of strain FML-4T revealed a genome size of 7.3 Mbp and a G+C content of 63.0 mol%. Based on the results obtained by phylogenetic and chemotaxonomic analyses, phenotypic characterization, average nucleotide identity (ANI, similarity 77.3-75.4%), and digital DNA-DNA hybridization (dDDH, similarity 24.5-22.3%), it was concluded that strain FML-4T represented a novel species of the genus Rhizobium, for which the name Rhizobium flavescens sp. nov. was proposed (type strain FML-4T = CCTCC AB 2019354T = KCTC 62839T).

Analysis and modulation of aberration in an extreme ultraviolet lithography projector via rigorous simulation and a back propagation neural network.

Lens aberration is a critical factor affecting lithography, one that deteriorates the image fidelity and contrast. As the perfect lens does not exist, the aberration control is important for real optical systems, especially for extreme ultraviolet lithography (EUVL). By choosing the process variation band (PVB) and pattern shift (PS) as the lithographic performance indicators, the inverse analysis model for aberration control is proposed in this paper. First, the effects of aberration with 36 Zernike terms on lithography performance are forward analyzed. Using the definitive screening design (DSD) and with the help of statistical analysis methods of analysis of variance and F test, the combined Zernike terms leading to prominent PVB and PS are identified. After giving a brief introduction of backpropagation neural network (BPNN), the aberration control model based on DSD and BPNN is then established. Finally, several examples are analyzed to demonstrate the effectiveness and robustness of the aberration control model. Predicted results show that the optimum distribution of Zernike coefficients given by the aberration model can generate minimum impact on imaging quality, and this impact is very close to that of zero aberration. The results demonstrate that the BPNN-based aberration model has the potential to be an efficient guiding method for controlling the aberration of EUVL in the optical design stage.

Sphingomonas flavalba sp. nov., isolated from a procymidone-contaminated soil.

A novel Gram-stain-negative, strictly aerobic, non-spore-forming, motile with one polar flagellum and short rod-shaped bacterium, designated strain ZLT-5T, was isolated from procymidone-contaminated soil sampled in Nanjing, Jiangsu, PR China. Growth occurred at 26-37 °C (optimum, 37 °C), at pH 6.0-9.0 (pH 7.0) and in the presence of 0-1.5 % NaCl (0.5 %). Phylogenetic analysis based on the 16S rRNA gene sequences showed that strain ZLT-5T belonged to the genus Sphingomonas, with the highest sequence similarity to Sphingomonas kyeonggiensis THG-DT81T (96.6 %), followed by Sphingomonas dokdonensis DSM 21029T (96.5 %) and Sphingomonas silvisoli RP18T (96.3 %). The G+C content of strain ZLT-5T was 68.0 mol% (draft genome sequence). The average nucleotide identity value of the draft genomes between strain ZLT-5T and S. kyeonggiensis THG-DT81T was 75.4 %. Strain ZLT-5T contained ubiquinone-10 as the predominant isoprenoid quinone and sym-homospermidine as the major polyamine. The major polar lipids consisted of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylcholine, phosphoaminolipid and sphingoglycolipid. The main cellular fatty acids (>10 % of the total fatty acids) of strain ZLT-5T were C17 : 1ω6c, summed feature 3 (C16 : 1ω7c and/or C15 : 0 ISO 2-OH) and summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c). Based on phylogenetic analysis and physiological and biochemical characterization, strain ZLT-5T is described as a novel species of the genus Sphingomonas, for which the name Sphingomonas flavalba sp. nov. is proposed. The type strain is ZLT-5T (=CCTCC AB 2018188T=KCTC 62840T).

Spirosoma sordidisoli sp. nov., a propanil-degrading bacterium isolated from a herbicide-contaminated soil.

A novel Gram-stain negative, aerobic, rod-shaped, asporogenous, propanil-degrading bacterial strain, TY50T, was isolated from a herbicide-contaminated soil in Nanjing, China. Strain TY50T was found to grow optimally at pH 9.0, 30 °C and in the absence of NaCl. The G + C content of the total DNA was determined to be 55.5 mol%. The 16S rRNA gene sequence of strain TY50T shows high identity to that of Spirosoma lacussanchae CPCC 100624T (99.3%), Spirosoma metallicum PR1014kT (94.8%) and Spirosoma soli MIMBbqt12T (94.6%). DNA-DNA hybridization indicated that the isolate had relatively low levels of DNA-DNA relatedness with S. lacussanchae CPCC 100624T (48.3%). Average nucleotide identity and the digital DNA-DNA hybridizations for draft genomes between strain TY50T and S. lacussanchae CPCC 100624T were 93.2% and 51.0%, respectively. The major cellular fatty acids of strain TY50T were identified as C16:1ω5c (24.5%) and summed feature 3 (C16:1ω6c/C16:1ω7c, 40.7%). MK-7 was found to be the predominant respiratory quinone. The major polar lipid profile includes phosphatidylethanolamine, an unidentified lipid and an unidentified aminolipid. These chemotaxonomic data support the affiliation of strain TY50T with the members of the genus Spirosoma. Strain TY50T can be distinguished from its close phylogenetic neighbours based on its phenotypic, genotypic, and chemotaxonomic features. Therefore, strain TY50T represents a novel member of the genus Spirosoma, for which the name Spirosoma sordidisoli sp. nov. is proposed. The type strain is TY50T (= KCTC 62494T = CCTCC AB 2018041T).

Mitigating the influence of wafer topography on the implantation process in optical lithography.

Residual resist in the gate caused by wafer topography would prevent part of the fins from ion implantation. It is a big concern in semiconductor manufacturing. The optical interference intensity improvement of the region located between two gates induced by substituting the material of the sidewall of the gate with silicon oxide is discussed in this Letter. The relationship between the thickness of silicon oxide and optical intensity at the fin's top is also established from the rigorous coupled wave analysis method. Based on this correlation, the method for mitigating the wafer topography of the implantation process is put forward and evaluated from rigorous numerical simulations. The proposed method requires neither a specific system setup nor an additional etch process, which is a tremendous cost saver in mass production.

Optimal chin-brow vertical angle for sagittal visual fields in ankylosing spondylitis kyphosis.

PURPOSE: Chin-brow vertical angle (CBVA) is very important in correction of thoracolumbar kyphotic deformity in ankylosing spondylitis (AS), especially for the patients with cervical ankylosis. In previous study, Suk et al. stated that the patients with CBVA between -10° and 10° had better horizontal gaze. Unfortunately, in our clinical practice, we found the patients with CBVA between -10° and 10° after surgery usually complained of difficulty in cooking, cleaning, desk working and the like, although they had excellent horizontal gaze. In other words, for the patients with cervical ankylosis, good horizontal gaze existed together with poor downward gaze. Then, which condition do the patients prefer? Is there a compromise solution that makes a better quality life possible for the patients? In this research, we studied AS patients with cervical ankylosis, aiming to investigate the optimal CBVA for deformity correction. METHODS: 25 AS thoracolumbar kyphotic patients with cervical ankylosis were studied, whose function and expectation of visual field related to life quality were assessed by questionnaire before and after surgery. Pre- and post-operative CBVA were obtained on lateral photos of the patients with free-standing posture, and 50 cases of CBVA were included, which were divided into six groups according to the angle irrespective of surgery (Group A, CBVA <0°; Group B, 0° ≤ CBVA < 10°; Group C, 10° ≤ CBVA < 20°; Group D, 20° ≤ CBVA < 30°; Group E, 30° ≤ CBVA < 40°; Group F, CBVA ≥ 40°). Kruskal-Wallis test was used to assess all the groups in terms of various items in the questionnaire, while Mann-Whitney test was used to assess every two groups. RESULTS: In overall evaluation, Group C (10°-20°) obtained the optimal expectation (p < 0.05); Group B, C and D (0°-30°) obtained better function (p < 0.05), and there was no significant difference between the 3 groups. In appearance, Group A, B and C (<20°) were better than the other groups both in function and expectation (p < 0.05), without dramatic difference among the three groups. In outdoor activities, Group A, B, C and D (<30°) were better in most of the items (p < 0.05). In indoor activities, Group C and D (10-30°) were much better (p < 0.05). CONCLUSION: AS thoracolumbar kyphotic patients with cervical ankylosis had the best satisfaction when 10° ≤ CBVA < 20°.

Exploration of factors affecting hemodynamic stability following pheochromocytoma resection - cohort study.

Purpose: Surgery is the only way to cure pheochromocytoma; however, postoperative hemodynamic instability is one of the main causes of serious complications and even death. This study's findings provide some guidance for improved clinical management. Patients and methods: This study was to investigate the factors leading to postoperative hemodynamic instability in the postoperative pathology indicated pheochromocytoma from May 2016 to May 2022. They were divided into two groups according to whether vasoactive drugs were used for a median number of days or more postoperatively. The factors affecting the postoperative hemodynamics in the perioperative period (preoperative, intraoperative, and postoperative) were then evaluated. Results: The median number of days requiring vasoactive drug support postoperatively was three in 234 patients, while 118 (50.4%) patients required vasoactive drug support for three days or more postoperatively. The results of the multivariate analysis indicated more preoperative colloid use (odds ratio [OR]=1.834, confidence interval [CI]:1.265-2.659, P=0.001), intraoperative use of vasoactive drug (OR=4.174, CI:1.882-9.258, P<0.001), and more postoperative crystalloid solution input per unit of body weight per day (ml/kg/d) (OR=1.087, CI:1.062-1.112, P<0.001) were risk factors for predicting postoperative hemodynamic instability. The optimal cutoff point of postoperative crystalloid use were 42.37 ml/kg/d. Conclusion: Hemodynamic instability is a key issue for consideration in the perioperative period of pheochromocytoma. The amount of preoperative colloid use, the need for intraoperative vasoactive drugs, and postoperative crystalloid solution are risk factors for predicting postoperative hemodynamic instability (registration number: ChiCT2300071166).

Capture of Group A Streptococcus by Open-Microfluidic CandyCollect Device in Pediatric Patients.

Importance: Obtaining high-quality samples to diagnose streptococcal pharyngitis in pediatric patients is challenging due to discomfort associated with traditional pharyngeal swabs. This may cause reluctance to go to the clinic, inaccurate diagnosis, or inappropriate treatment for children with sore throat. Objective: Determine the efficacy of using CandyCollect, a lollipop-inspired open-microfluidic pathogen collection device, to capture Group A Streptococcus (GAS) and compare user preference for CandyCollect, conventional pharyngeal swabs, or mouth swabs among children with pharyngitis and their caregivers. Design: Participants of this cohort study were recruited over a 7-month period in 2022 - 2023. Setting: This study was conducted at an ambulatory care clinic that serves pediatric patients in the Madison, Wisconsin, metropolitan area. Participants: Study participants were diagnosed with GAS pharyngitis using a traditional pharyngeal swab via rapid antigen detection test (RADT); those testing positive were approached or reached out to about participation in the study. A total of 74 caregiver/children dyads were contacted about the study: 23 declined to participate; 21 were not eligible; and 30 willing and eligible participants were admitted into the study. A caregiver provided verbal consent and parental permission, and all children provided verbal assent. Immediately after the standard of care visit in which the throat swab was obtained, a research nurse guided participants through collecting oral samples: CandyCollect device and mouth swab (ESwab TM ). CandyCollect and mouth swab samples were analyzed for GAS by quantitative polymerase chain reaction (qPCR) at the University of Washington. Exposure: Detection of salivary GAS using qPCR analysis of samples obtained from CandyCollect devices and mouth swabs. Main Outcomes and Measures: The proportion of pediatric patients with GAS pharyngitis, as determined by a positive pharyngeal swab tested via a RADT, who were also positive using a CandyCollect and mouth swab analyzed by qPCR. Results: All child participants (30/30) were positive for GAS by qPCR on both the mouth swab and CandyCollect. Caregivers ranked CandyCollect as a good sampling method overall (27/30), and all caregivers (30/30) would recommend the CandyCollect for children 5 years and older. Twenty-three of 30 children "really like" the taste and 24/30 would prefer to use the CandyCollect if a future test was needed. All caregivers (30/30) and most children (28/30) would be willing to use the CandyCollect device at home. Conclusion and relevance: All participants tested positive for GAS on all three collection methods (pharyngeal swab, mouth swab, and CandyCollect). While both caregivers and children like the CandyCollect device, some caregivers would prefer a shorter collection time. Future work includes additional studies with larger cohorts presenting with pharyngitis of unknown etiology and shortening collection time, while maintaining the attractive form of the device. Trial Registration: Registry name: ClinicalTrials.gov ClinicalTrials.gov Identifier: NCT05175196 Weblink: https://classic.clinicaltrials.gov/ct2/show/NCT05175196. Key Points: Question: In pediatric patients with Group A Streptococcus pharyngitis, how do test results and user experience compare across three sampling methods-CandyCollect devices, mouth swabs, and pharyngeal swabs?Findings: In this cohort study of 30 children, aged 5-14 years, saliva samples were collected with CandyCollect devices and mouth swabs and analyzed via qPCR. The results show CandyCollect, a pathogen collection tool preferred by children, had 100% concordance with the results from pharyngeal swabs positive with a rapid antigen detection test performed as part of their clinical care.Meaning: With further development and testing, the CandyCollect device may potentially become an alternative sampling tool for the diagnosis of streptococcal pharyngitis.

Effect of microculture on cell metabolism and biochemistry: do cells get stressed in microchannels?

Microfluidics is emerging as a promising platform for cell culture, enabling increased microenvironment control and potential for integrated analysis compared to conventional macroculture systems such as well plates and Petri dishes. To advance the use of microfluidic devices for cell culture, it is necessary to better understand how miniaturization affects cell behavior. In particular, microfluidic devices have significantly higher surface-area-to-volume ratios than conventional platforms, resulting in lower volumes of media per cell, which can lead to cell stress. We investigated cell stress under a variety of culture conditions using three cell lines: parental HEK (human embryonic kidney) cells and transfected HEK cells that stably express wild-type (WT) and mutant (G601S) human ether-a-go-go related gene (hERG) potassium channel protein. These three cell lines provide a unique model system through which to study cell-type-specific responses in microculture because mutant hERG is known to be sensitive to environmental conditions, making its expression a particularly sensitive readout through which to compare macro- and microculture. While expression of WT-hERG was similar in microchannel and well culture, the expression of mutant G601S-hERG was reduced in microchannels. Expression of the endoplasmic reticulum (ER) stress marker immunoglobulin binding protein (BiP) was upregulated in all three cell lines in microculture. Using BiP expression, glucose consumption, and lactate accumulation as readouts we developed methods for reducing ER stress including properly increasing the frequency of media replacement, reducing cell seeding density, and adjusting the serum concentration and buffering capacity of culture medium. Indeed, increasing the buffering capacity of culture medium or frequency of media replacement partially restored the expression of the G601S-hERG in microculture. This work illuminates how biochemical properties of cells differ in macro- and microculture and suggests strategies that can be used to modify cell culture protocols for future studies involving miniaturized culture platforms.

Involvement of Mrgprd-expressing nociceptors-recruited spinal mechanisms in nerve injury-induced mechanical allodynia.

Mechanical allodynia and hyperalgesia are intractable symptoms lacking effective clinical treatments in patients with neuropathic pain. However, whether and how mechanically responsive non-peptidergic nociceptors are involved remains elusive. Here, we showed that von Frey-evoked static allodynia and aversion, along with mechanical hyperalgesia after spared nerve injury (SNI) were reduced by ablation of MrgprdCreERT2-marked neurons. Electrophysiological recordings revealed that SNI-opened Aß-fiber inputs to laminae I-IIo and vIIi, as well as C-fiber inputs to vIIi, were all attenuated in Mrgprd-ablated mice. In addition, priming chemogenetic or optogenetic activation of Mrgprd+ neurons drove mechanical allodynia and aversion to low-threshold mechanical stimuli, along with mechanical hyperalgesia. Mechanistically, gated Aß and C inputs to vIIi were opened, potentially via central sensitization by dampening potassium currents. Altogether, we uncovered the involvement of Mrgprd+ nociceptors in nerve injury-induced mechanical pain and dissected the underlying spinal mechanisms, thus providing insights into potential therapeutic targets for pain management.

A glutamatergic DRN-VTA pathway modulates neuropathic pain and comorbid anhedonia-like behavior in mice.

Chronic pain causes both physical suffering and comorbid mental symptoms such as anhedonia. However, the neural circuits and molecular mechanisms underlying these maladaptive behaviors remain elusive. Here using a mouse model, we report a pathway from vesicular glutamate transporter 3 neurons in the dorsal raphe nucleus to dopamine neurons in the ventral tegmental area (VGluT3DRN→DAVTA) wherein population-level activity in response to innocuous mechanical stimuli and sucrose consumption is inhibited by chronic neuropathic pain. Mechanistically, neuropathic pain dampens VGluT3DRN → DAVTA glutamatergic transmission and DAVTA neural excitability. VGluT3DRN → DAVTA activation alleviates neuropathic pain and comorbid anhedonia-like behavior (CAB) by releasing glutamate, which subsequently promotes DA release in the nucleus accumbens medial shell (NAcMed) and produces analgesic and anti-anhedonia effects via D2 and D1 receptors, respectively. In addition, VGluT3DRN → DAVTA inhibition produces pain-like reflexive hypersensitivity and anhedonia-like behavior in intact mice. These findings reveal a crucial role for VGluT3DRN → DAVTA → D2/D1NAcMed pathway in establishing and modulating chronic pain and CAB.

At-home saliva sampling in healthy adults using CandyCollect, a lollipop-inspired device.

Respiratory infections are common in children, and there is a need for user-friendly collection methods. Here, we performed the first human subjects study using the CandyCollect device, a lollipop inspired saliva collection device. 1 We showed the CandyCollect device can be used to collect salivary bacteria from healthy adults using Streptococcus mutans and Staphylococcus aureus as proof-of-concept commensal bacteria. We enrolled healthy adults in a nationwide (USA) remote study in which participants were sent study packages containing CandyCollect devices and traditional commercially available oral swabs and spit tubes. Participants sampled themselves at home, completed usability and user preference surveys, and mailed the samples back to our laboratory for analysis by qPCR. Our results showed that for participants in which a given bacterium ( S. mutans or S. aureus ) was detected in one or both of the commercially available methods (oral swab and/or spit tubes), CandyCollect devices had a 100% concordance with the positive result (n=14 participants). Furthermore, the CandyCollect device was ranked the highest preference sampling method among the three sampling methods by 26 participants surveyed (combining survey results across two enrollment groups). We also showed that the CandyCollect device has a shelf life of up to 1 year at room temperature, a storage period that is convenient for clinics or patients to keep the CandyCollect device and use it any time. Taken together, we have demonstrated that the CandyCollect is a user-friendly saliva collection tool that has the potential to be incorporated into diagnostic assays in clinic visits and telemedicine.

Protonated cross-linkable nanocomposite coatings with outstanding underwater superoleophobic and anti-viscous oil-fouling properties for crude oil/water separation.

Superhydrophilic/underwater superoleophobic coatings that effectively prevent viscous oil contamination have been of considerable interest for the great potential in oil spill remediation and oilfield wastewater treatment. In the present work, a protonated cross-linkable nanocomposite coating with robust underwater superoleophobicity and intensified hydration capability is proposed through the synthesis of active polymeric nanocomplex (PNC), cross-linking reaction between PNC and hydrophilic chitosan (CS), and final protonation to further improve water affinity. Benefiting from the hierarchical structure and strong hydration capability induced by electrostatic interactions and hydrogen bondings, the nanocomposite coating coated textile exhibits excellent superhydrophilicity (within 0.28 s with water contact angle reaching 0°), underwater superoleophobicity (underwater crude oil contact angle at 160°), and ultralow oil adhesion even to highly viscous silicone oil. Moreover, the nanocomposite coating presents a robust chemical resistance, mechanical tolerance, and storage stability. Simultaneously, the nanocomposite coating adapts well to various porous substrates (e.g., stainless steel mesh and Ni sponge) with great anti-oil-fouling and self-cleaning performances. Importantly, the coating coated textile is successfully applied in crude oil/water separation with excellent efficiency and repeatability. The findings conceivably stand out as a new methodology to fabricate superhydrophilic/underwater superoleophobic materials with outstanding anti-viscous oil-fouling property for practically treating oily wastewater.

The T-Type Calcium Channel Cav3.2 in Somatostatin Interneurons in Spinal Dorsal Horn Participates in Mechanosensation and Mechanical Allodynia in Mice.

Somatostatin-positive (SOM+) neurons have been proposed as one of the key populations of excitatory interneurons in the spinal dorsal horn involved in mechanical pain. However, the molecular mechanism for their role in pain modulation remains unknown. Here, we showed that the T-type calcium channel Cav3.2 was highly expressed in spinal SOM+ interneurons. Colocalization of Cacna1h (which codes for Cav3.2) and SOM tdTomato was observed in the in situ hybridization studies. Fluorescence-activated cell sorting of SOM tdTomato cells in spinal dorsal horn also proved a high expression of Cacna1h in SOM+ neurons. Behaviorally, virus-mediated knockdown of Cacna1h in spinal SOM+ neurons reduced the sensitivity to light touch and responsiveness to noxious mechanical stimuli in naïve mice. Furthermore, knockdown of Cacna1h in spinal SOM+ neurons attenuated thermal hyperalgesia and dynamic allodynia in the complete Freund's adjuvant-induced inflammatory pain model, and reduced both dynamic and static allodynia in a neuropathic pain model of spared nerve injury. Mechanistically, a decrease in the percentage of neurons with Aß-eEPSCs and Aß-eAPs in superficial dorsal horn was observed after Cacna1h knockdown in spinal SOM+ neurons. Altogether, our results proved a crucial role of Cav3.2 in spinal SOM+ neurons in mechanosensation under basal conditions and in mechanical allodynia under pathological pain conditions. This work reveals a molecular basis for SOM+ neurons in transmitting mechanical pain and shows a functional role of Cav3.2 in tactile and pain processing at the level of spinal cord in addition to its well-established peripheral role.

Parallel Spinal Pathways for Transmitting Reflexive and Affective Dimensions of Nocifensive Behaviors Evoked by Selective Activation of the Mas-Related G Protein-Coupled Receptor D-Positive and Transient Receptor Potential Vanilloid 1-Positive Subsets of Nociceptors.

The high incidence of treatment-resistant pain calls for the urgent preclinical translation of new analgesics. Understanding the behavioral readout of pain in animals is crucial for efficacy evaluation when developing novel analgesics. Mas-related G protein-coupled receptor D-positive (Mrgprd+) and transient receptor potential vanilloid 1-positive (TRPV1+) sensory neurons are two major non-overlapping subpopulations of C-fiber nociceptors. Their activation has been reported to provoke diverse nocifensive behaviors. However, what kind of behavior reliably represents subjectively conscious pain perception needs to be revisited. Here, we generated transgenic mice in which Mrgprd+ or TRPV1+ sensory neurons specifically express channelrhodopsin-2 (ChR2). Under physiological conditions, optogenetic activation of hindpaw Mrgprd+ afferents evoked reflexive behaviors (lifting, etc.), but failed to produce aversion. In contrast, TRPV1+ afferents activation evoked marked reflexive behaviors and affective responses (licking, etc.), as well as robust aversion. Under neuropathic pain conditions induced by spared nerve injury (SNI), affective behaviors and avoidance can be elicited by Mrgprd+ afferents excitation. Mechanistically, spinal cord-lateral parabrachial nucleus (lPBN) projecting neurons in superficial layers (lamina I-II o ) were activated by TRPV1+ nociceptors in naïve conditions or by Mrgprd+ nociceptors after SNI, whereas only deep spinal cord neurons were activated by Mrgprd+ nociceptors in naïve conditions. Moreover, the excitatory inputs from Mrgprd+ afferents to neurons within inner lamina II (II i ) are partially gated under normal conditions. Altogether, we conclude that optogenetic activation of the adult Mrgprd+ nociceptors drives non-pain-like reflexive behaviors via the deep spinal cord pathway under physiological conditions and drives pain-like affective behaviors via superficial spinal cord pathway under pathological conditions. The distinct spinal pathway transmitting different forms of nocifensive behaviors provides different therapeutic targets. Moreover, this study appeals to the rational evaluation of preclinical analgesic efficacy by using comprehensive and suitable behavioral assays, as well as by assessing neural activity in the two distinct pathways.

An open microfluidic coculture model of fibroblasts and eosinophils to investigate mechanisms of airway inflammation.

Interactions between fibroblasts and immune cells play an important role in tissue inflammation. Previous studies have found that eosinophils activated with interleukin-3 (IL-3) degranulate on aggregated immunoglobulin G (IgG) and release mediators that activate fibroblasts in the lung. However, these studies were done with eosinophil-conditioned media that have the capacity to investigate only one-way signaling from eosinophils to fibroblasts. Here, we demonstrate a coculture model of primary normal human lung fibroblasts (HLFs) and human blood eosinophils from patients with allergy and asthma using an open microfluidic coculture device. In our device, the two types of cells can communicate via two-way soluble factor signaling in the shared media while being physically separated by a half wall. Initially, we assessed the level of eosinophil degranulation by their release of eosinophil-derived neurotoxin (EDN). Next, we analyzed the inflammation-associated genes and soluble factors using reverse transcription quantitative polymerase chain reaction (RT-qPCR) and multiplex immunoassays, respectively. Our results suggest an induction of a proinflammatory fibroblast phenotype of HLFs following the coculture with degranulating eosinophils, validating our previous findings. Additionally, we present a new result that indicate potential impacts of activated HLFs back on eosinophils. This open microfluidic coculture platform provides unique opportunities to investigate the intercellular signaling between the two cell types and their roles in airway inflammation and remodeling.