Ultrafast Microfluidic Immunoassays Towards Real-time Intervention of Cytokine Storms

Biomarker-guided precision medicine holds great promise to provide personalized therapy with a good understanding of the molecular or cellular data of an individual patient. However, implementing this approach in critical care uniquely faces enormous challenges as it requires obtaining “real-time” data with high sensitivity, reliability, and multiplex capacity near the patient’s bedside in the quickly evolving illness. Current immunodiagnostic platforms generally compromise assay sensitivity and specificity for speed or face significantly increased complexity and cost for highly multiplexed detection with low sample volume. This thesis introduces two novel ultrafast immunoassay platforms: one is a machine learning-based digital molecular counting assay, and the other is a label-free nano-plasmonic sensor integrated with an electrokinetic mixer. Both of them incorporate microfluidic approaches to pave the way for near-real-time interventions of cytokine storms. In the first part of the thesis, we present an innovative concept and the theoretical study that enables ultrafast measurement of multiple protein biomarkers (<1 min assay incubation) with comparable sensitivity to the gold standard ELISA method. The approach, which we term “pre-equilibrium digital enzyme-linked immunosorbent assay” (PEdELISA) incorporates the single-molecular counting of proteins at the early, pre-equilibrium state to achieve the combination of high speed and sensitivity. We experimentally demonstrated the assay’s application in near-real-time monitoring of patients receiving chimeric antigen receptor (CAR) T-cell therapy and for longitudinal serum cytokine measurements in a mouse sepsis model. In the second part, we report the further development of a machine learning-based PEdELISA microarray data analysis approach with a significantly extended multiplex capacity using the spatial-spectral microfluidic encoding technique. This unique approach, together with a convolutional neural network-based image analysis algorithm, remarkably reduced errors faced by the highly multiplexed digital immunoassay at low analyte concentrations. As a result, we demonstrated the longitudinal data collection of 14 serum cytokines in human patients receiving CAR-T cell therapy at concentrations < 10pg/mL with a sample volume < 10 µL and 5-min assay incubation. In the third part, we demonstrate the clinical application of a machine learning-based digital protein microarray platform for rapid multiplex quantification of cytokines from critically ill COVID-19 patients admitted to the intensive care unit. The platform comprises two low-cost modules: (i) a semi-automated fluidic dispensing module that can be operated inside a biosafety cabinet to minimize the exposure of technician to the virus infection and (ii) a compact fluorescence optical scanner for the potential near-bedside readout. The automated system has achieved high interassay precision (~10% CV) with high sensitivity (<0.4pg/mL). Our data revealed large subject-to-subject variability in patient responses to anti-inflammatory treatment for COVID-19, reaffirming the need for a personalized strategy guided by rapid cytokine assays. Lastly, an AC electroosmosis-enhanced localized surface plasmon resonance (ACE-LSPR) biosensing device was presented for rapid analysis of cytokine IL-1β among sepsis patients. The ACE-LSPR device is constructed using both bottom-up and top-down sensor fabrication methods, allowing the seamless integration of antibody-conjugated gold nanorod (AuNR) biosensor arrays with microelectrodes on the same microfluidic platform. Applying an AC voltage to microelectrodes while scanning the scattering light intensity variation of the AuNR biosensors results in significantly enhanced biosensing performance. The technologies developed have enabled new capabilities with broad application to advance precision medicine of life-threatening acute illnesses in critical care, which potentially will allow the clinical team to make individualized treatment decisions based on a set of time-resolved biomarker signatures.PHDMechanical EngineeringUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/163129/1/yujing_1.pd

Single Image Super-Resolution

Super-Resolution (SR) of a single image is a classic problem in computer vision. The goal of image super-resolution is to produce a high-resolution image from a low-resolution image. This paper presents a popular model, super-resolution convolutional neural network (SRCNN), to solve this problem. This paper also examines an improvement to SRCNN using a methodology known as generative adversarial net- work (GAN) which is better at adding texture details to the high resolution output

The synthesis and characterization of a xanthan gum-acrylamide-trimethylolpropane triglycidyl ether hydrogel

peer-reviewedTo improve the thermal stability and adsorption performance, xanthan gum was modified with acrylamide and trimethylolpropane triglycidyl ether (TTE). The modified xanthan gum (XGTTE) was characterized by Fourier transform infrared (FT-IR) spectroscopy, X-ray diffractogram (XRD), differential scanning calorimetry (DSC) and scanning electron microscopy (SEM). The characteristic peaks at 3449, 1655, 1611 and 1420 cm−1 in the FT-IR confirm the modification. The XGTTE crystal grew well upon addition of TTE. The XRD and DSC data revealed that the XGTTE enhanced its thermal stability. Analysis of SEM revealed that the grafting introduced major changes on the microstructure making it porous and resulting in the adsorption of crystal violet (CV) with flocculation. The CV adsorption capacity of the hydrogel with different dosages of TTE (XGTTE2, XGTTE3, XGTTE4, XGTTE5 and XGTTE6) were between 28.13 with 35.12 mg/g. In addition, the adsorption capacity, thermal stability, and swelling property of XGTTE4 were the best

Perfluorocarbon restrains inflammation and cell apoptosis in rats with lung ischemia reperfusion injury via down regulation of TLR4/NF κB signaling pathway

Purpose: To exaluate the effect of perfluorocarbon on lung ischemia-reperfusion injury in rats, and to unravel the potential underlying mechanism.Methods: A total of 36 Sprague-Dawley (SD) rats were randomly assigned to sham group, model group, and perfluorocarbon group (12 rats per group). The levels of inflammatory factors (TNF-α and IL1β) were determined using enzyme-linked immunosorbent assay (ELISA). Cell apoptosis was evaluated by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay, while Western blotting was conducted to determine the protein expressions of TLR4 and NF-κB.Results: The levels of inflammatory factors in the model and perfluorocarbon groups were significantly higher than those in operation group (p &lt; 0.05), while their levels in perfluorocarbon group were significantly lower than in model group (p &lt; 0.05). The mRNA expression levels of TNF-α and IL-1β in lung tissues rose significantly in both model and perfluorocarbon groups when compared with those in sham group (p &lt; 0.05), but declined significantly in the perfluorocarbon group in comparison with those in model group (p &lt; 0.05). Furthermore, the perfluorocarbon group exhibited a significantly lower cell apoptosis than model group (p &lt; 0.05). The relative protein expression levels of TLR4 and NF-κBdeclined significantly in perfluorocarbon group than in model group.Conclusions: Perfluorocarbon down-regulates the TLR4/NF-κB signaling pathway, and inhibits inflammation a

An Integrated Plasmo‐Photoelectronic Nanostructure Biosensor Detects an Infection Biomarker Accompanying Cell Death in Neutrophils

Bacterial infections leading to sepsis are a major cause of deaths in the intensive care unit. Unfortunately, no effective methods are available to capture the early onset of infectious sepsis near the patient with both speed and sensitivity required for timely clinical treatment. To fill the gap, the authors develop a highly miniaturized (2.5 × 2.5 µm2) plasmo‐photoelectronic nanostructure device that detected citrullinated histone H3 (CitH3), a biomarker released to the blood circulatory system by neutrophils. Rapidly detecting CitH3 with high sensitivity has the great potential to prevent infections from developing life‐threatening septic shock. To this end, the author’s device incorporates structurally engineered arrayed hemispherical gold nanoparticles that are functionalized with high‐affinity antibodies. A nanoplasmonic resonance shift induces a photoconduction increase in a few‐layer molybdenum disulfide (MoS2) channel, and it provides the sensor signal. The device achieves label‐free detection of serum CitH3 with a 5‐log dynamic range from 10−4 to 101 ng mL and a sample‐to‐answer time <20 min. Using this biosensor, the authors longitudinally measure the dynamic CitH3 profiles of individual living mice in a sepsis model at high resolution over 12 hours. The developed biosensor may be poised for future translation to personalized management of systemic bacterial infections.The lack of an appropriate biosensing technology to detect the early onset of bacterial infections has prohibited timely clinical treatment of sepitc shock. This article presents a highly miniaturized plasmo‐photoelectronic device incorporating high‐affinity antibody‐conjugated hemispherical gold nanoparticles and a few‐layer molybdenum disulfide (MoS2) photoconductive channel to detect a blood biomarker released by neutrophils with high speed and sensitivity.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/152883/1/smll201905611-sup-0001-SuppMat.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/152883/2/smll201905611_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/152883/3/smll201905611.pd

Shufeng Jiedu Capsules Alleviate Lipopolysaccharide-Induced Acute Lung Inflammatory Injury via Activation of GPR18 by Verbenalin

Background/Aims: Acute respiratory tract infection (ARTI) is the most common reason for outpatient physician office visits. Although powerful and significant in the treatment of infections, antibiotics used for ARTI inappropriately have been an important contributor to antibiotic resistance. We previously reported that Shufeng Jiedu Capsule (SJC) can effectively amplify anti-inflammatory signaling during infection. In this study, we aimed to systematically explore its composition and the mechanism of its effects in ARTI. Methods: Pseudomonas aeruginosa (PAK) strain was used to generate a mouse model of ARTI, which were then treated with different drugs or compounds to determine the corresponding anti-inflammatory roles. High-performance liquid chromatography-quadrupole time of flight-tandem mass spectrometry. was conducted to detect the chemical compounds in SJC. RNAs from the lung tissues of mice were prepared for microarray analysis to reveal globally altered genes and the pathways involved after SJC treatment. Results: SJC significantly inhibited the expression and secretion of inflammatory factors from PAK-induced mouse lung tissues or lipopolysaccharide-induced peritoneal macrophages. Verbenalin, one of the bioactive compounds identified in SJC, also showed notable anti-inflammatory effects. Microarray data revealed numerous differentially expressed genes among the different treatment groups; here, we focused on studying the role of GPR18. We found that the anti-inflammatory role of verbenalin was attenuated in GPR18 knockout mice compared with wild-type mice, although no statistically significant difference was observed in the untreated PAK-induced mice types. Conclusion: Our data not only showed the chemical composition of SJC, but also demonstrated that verbenalin was a significant anti-inflammatory compound, which may function through GPR18

A novel DSPP mutation causes dentinogenesis imperfecta type II in a large Mongolian family

<p>Abstract</p> <p>Background</p> <p>Several studies have shown that the clinical phenotypes of dentinogenesis imperfecta type II (DGI-II) may be caused by mutations in <it>dentin sialophosphoprotein </it>(<it>DSPP</it>). However, no previous studies have documented the clinical phenotype and genetic basis of DGI-II in a Mongolian family from China.</p> <p>Methods</p> <p>We identified a large five-generation Mongolian family from China with DGI-II, comprising 64 living family members of whom 22 were affected. Linkage analysis of five polymorphic markers flanking <it>DSPP </it>gene was used to genotype the families and to construct the haplotypes of these families. All five DSPP exons including the intron-exon boundaries were PCR-amplified and sequenced in 48 members of this large family.</p> <p>Results</p> <p>All affected individuals showed discoloration and severe attrition of their teeth, with obliterated pulp chambers and without progressive high frequency hearing loss or skeletal abnormalities. No recombination was found at five polymorphic markers flanking DSPP in the family. Direct DNA sequencing identified a novel A→G transition mutation adjacent to the donor splicing site within intron 3 in all affected individuals but not in the unaffected family members and 50 unrelated Mongolian individuals.</p> <p>Conclusion</p> <p>This study identified a novel mutation (IVS3+3A→G) in <it>DSPP</it>, which caused DGI-II in a large Mongolian family. This expands the spectrum of mutations leading to DGI-II.</p

Estimation of effect of voids on frequency response of mountain tunnel lining based on microtremor method

Nowadays, aged tunnels keep continuously increasing all over the world, which require effective inspection methods to assess their health conditions. In this study, both in situ acceleration wave measurements and numerical simulations were carried out to study the microtremor characteristics of mountain tunnel lining. Power spectrum density (PSD) of signals was calculated and peak frequencies were identified using the peak-picking method. Discontinuous contacts between rock masses and lining concrete were simplified as weak interfaces with low stiffness, which play the role of elastic supporting during dynamic calculation. Influences of voids, rock type and concrete type on their peak frequencies were evaluated. The results of the numerical analysis show that the normal stiffness of rock-concrete interface has strong correlation with the magnitude of peak frequency. The frequency response of tunnel lining is affected by the presence of voids located around tunnel circumference behind lining. The first peak frequency is dependent on the void size and location. The larger the void size and location angle, the greater the decrease of the first peak frequency. The peak frequency also decreases as the strength of concrete decreases, and is not affected by the change of properties of unweathered rock masses. Additional frequency modes can be identified when voids are not located on the central axis of tunnel. The first peak frequency variation can be considered to be intimately linked with the stress state of the tunnel lining influenced by the existence of voids