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A surgical method in mice can selectively remove dorsal root ganglia (DRG) at specific spinal levels without interfering with other nerves, providing insight on thoracolumbar vs. lumbosacral DRG contributions to pain signalling and behaviour.

Abstract Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for the infectious disease COVID-19, which has rapidly become an international pandemic with significant impact on healthcare systems and the global economy. To assist antiviral therapy and vaccine development efforts, we performed a natural history/time course study of SARS-CoV-2 infection in ferrets to characterise and assess the suitability of this animal model. Ten ferrets of each sex were challenged intranasally with 4.64 × 104 TCID50 of SARS-CoV-2 isolate Australia/VIC01/2020 and monitored for clinical disease signs, viral shedding, and tissues collected post-mortem for histopathological and virological assessment at set intervals. We found that SARS-CoV-2 replicated in the upper respiratory tract of ferrets with consistent viral shedding in nasal wash samples and oral swab samples up until day 9. Infectious SARS-CoV-2 was recovered from nasal washes, oral swabs, nasal turbinates, pharynx, and olfactory bulb samples within 3–7 days post-challenge; however, only viral RNA was detected by qRT-PCR in samples collected from the trachea, lung, and parts of the gastrointestinal tract. Viral antigen was seen exclusively in nasal epithelium and associated sloughed cells and draining lymph nodes upon immunohistochemical staining. Due to the absence of clinical signs after viral challenge, our ferret model is appropriate for studying asymptomatic SARS-CoV-2 infections and most suitable for use in vaccine efficacy studies.

Nick Spencer et al. made simultaneous multi-site electrophysiological recordings with video imaging of colonic wall movements from ex vivo mouse colon, in order to correlate propulsion of content with underlying electrical signals from the smooth muscle. Their results demonstrate that excitatory and inhibitory junction potentials are synchronized in both the proximal and distal colon, suggesting that the enteric nervous system network communicates over a longer range than previously expected.

Sensory stimuli from the uterus are detected by spinal afferent neurons whose cell bodies arise from thoracolumbar and lumbosacral dorsal root ganglia (DRG). Using an in vivo survival surgical technique developed in our laboratory to remove select DRG from live mice, we recently quantified the topographical distribution of thoracolumbar spinal afferents innervating the mouse uterine horn, revealed by loss of immunoreactivity to calcitonin gene-related peptide (CGRP). Here, we used the same technique to investigate the distribution of lumbosacral uterine spinal afferents, in which L5-S1 DRG were unilaterally removed from adult female C57BL/6J mice (N = 6). Following 10–12 days recovery, CGRP immunoreactivity was quantified along the length of uterine horns using fluorescence immunohistochemistry. Relative to myometrial thickness, overall CGRP density in uterine tissues ipsilateral to L5-S1 DRG removal was reduced compared to the DRG-intact, contralateral side (P = 0.0265). Regionally, however, myometrial CGRP density was unchanged in the cranial, mid, and caudal portions. Similarly, CGRP-expressing nerve fiber counts, network lengths, junctions, and the proportion of area occupied by CGRP immunoreactivity were unaffected by DRG removal (P ≥ 0.2438). Retrograde neuronal tracing from the caudal uterine horn revealed fewer spinal afferents here arise from lumbosacral than thoracolumbar DRG (P = 0.0442) (N = 4). These data indicate that, unlike thoracolumbar DRG, lumbosacral spinal afferent nerves supply relatively modest sensory innervation across the mouse uterine horn, with no regional specificity. We conclude most sensory information between the mouse uterine horn and central nervous system is likely relayed via thoracolumbar spinal afferents.

Abstract Vaccines against SARS-CoV-2 are likely to be critical in the management of the ongoing pandemic. A number of candidates are in Phase III human clinical trials, including ChAdOx1 nCoV-19 (AZD1222), a replication-deficient chimpanzee adenovirus-vectored vaccine candidate. In preclinical trials, the efficacy of ChAdOx1 nCoV-19 against SARS-CoV-2 challenge was evaluated in a ferret model of infection. Groups of ferrets received either prime-only or prime-boost administration of ChAdOx1 nCoV-19 via the intramuscular or intranasal route. All ChAdOx1 nCoV-19 administration combinations resulted in significant reductions in viral loads in nasal-wash and oral swab samples. No vaccine-associated adverse events were observed associated with the ChAdOx1 nCoV-19 candidate, with the data from this study suggesting it could be an effective and safe vaccine against COVID-19. Our study also indicates the potential for intranasal administration as a way to further improve the efficacy of this leading vaccine candidate.

How the enteric nervous system determines the pacing and propagation direction of neurogenic contractions along the colon remains largely unknown. We used a chemogenetic strategy to ablate enteric neurons expressing calretinin (CAL). Mice expressing human diphtheria toxin receptor (DTR) in CAL neurons were generated by crossing CAL-ires-Cre mice with Cre-dependent ROSA26-DTR mice. Immunohistochemical analysis revealed treatment with diphtheria toxin incurred a 42% reduction in counts of Hu-expressing colonic myenteric neurons (P = 0.036), and 57% loss of CAL neurons (comprising ∼25% of all Hu neurons; P = 0.004) compared to control. As proportions of Hu-expressing neurons, CAL neurons that contained nitric oxide synthase (NOS) were relatively spared (control: 15 ± 2%, CAL-DTR: 13 ± 1%; P = 0.145), while calretinin neurons lacking NOS were significantly reduced (control: 26 ± 2%, CAL-DTR: 18 ± 5%; P = 0.010). Colonic length and pellet sizes were significantly reduced without overt inflammation or changes in ganglionic density. Interestingly, colonic motor complexes (CMCs) persisted with increased frequency (mid-colon interval 111 ± 19 vs. 189 ± 24 s, CAL-DTR vs. control, respectively, P < 0.001), decreased contraction size (mid-colon AUC 26 ± 24 vs. 59 ± 13 gram/seconds, CAL-DTR vs. control, respectively, P < 0.001), and lacked preferential anterograde migration (P < 0.001). The functional effects of modest calretinin neuron ablation, particularly increased neurogenic motor activity frequencies, differ from models that incur general enteric neuron loss, and suggest calretinin neurons may contribute to pacing, force, and polarity of CMCs in the large bowel.

Acute aerobic exercise induces skeletal muscle mitochondrial gene expression, which in turn can increase muscle mitochondrial protein synthesis. In this regard, the peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α), is a master regulator of mitochondrial biogenesis, and thus mitochondrial protein synthesis. However, PGC-1α expression is impaired in muscle of humans with obesity in response to acute aerobic exercise. Therefore, we sought to determine whether muscle mitochondrial protein synthesis is also impaired under the same conditions in humans with obesity. To this end, we measured mitochondrial and mixed-muscle protein synthesis in skeletal muscle of untrained subjects with (body fat: 34.7 ± 2.3%) and without (body fat: 25.3 ± 3.3%) obesity in a basal period and during a continuous period that included a 45 min cycling exercise (performed at an intensity corresponding to 65% of heart rate reserve) and a 3-h post-exercise recovery. Exercise increased PGC-1α mRNA expression in muscle of subjects without obesity, but not in subjects with obesity. However, muscle mitochondrial protein synthesis did not increase in either subject group. Similarly, mixed-muscle protein synthesis did not increase in either group. Concentrations of plasma amino acids decreased post-exercise in the subjects without obesity, but not in the subjects with obesity. We conclude that neither mitochondrial nor mixed-muscle protein synthesis increase in muscle of humans during the course of a session of aerobic exercise and its recovery period in the fasting state irrespective of obesity.Trial Registration: The study has been registered within ClinicalTrials.gov (NCT01824173).

In electrophysiology, many methods have been proposed for the analysis of action potential firing frequencies. The aim of this study was to present an algorithm developed for a continuous wavelet transform that enables the filtering out of frequencies contributing to the shapes of action potentials (spikes), whilst retaining the frequencies that encode the periodicity of spike trains. The continuous wavelet transform allows us to decompose a signal into its constituent frequencies. A signal with a single event, such as a spike, is composed of frequencies that characterize the shape of the spike. A signal with two spikes will also be composed of frequencies characterizing the shape of the action potential, but in addition will include a substantial portion of its power at the frequency corresponding to the time-difference between the two spikes. This is achieved by clipping peaks from the wavelet amplitudes that are narrower than a given minimum number of phase cycles. We present some application examples in both synthetic signals and electrophysiological recordings. This new approach can provide a major new analytical tool for analysis of electrophysiological signals.

Small cell lung cancer (SCLC) patients frequently relapse and become resistant to chemotherapy. Here, the authors analyse the genomic and transcriptomic landscape of primary and relapsed SCLC patients as well as in vitro models, and discover that activation of WNT signalling can drive chemotherapy resistance.

Abstract Background Ever since the first atomic structure of an enzyme was solved, the discovery of the mechanism and dynamics of reactions catalyzed by biomolecules has been the key goal for the understanding of the molecular processes that drive life on earth. Despite a large number of successful methods for trapping reaction intermediates, the direct observation of an ongoing reaction has been possible only in rare and exceptional cases. Results Here, we demonstrate a general method for capturing enzyme catalysis “in action” by mix-and-inject serial crystallography (MISC). Specifically, we follow the catalytic reaction of the Mycobacterium tuberculosis β-lactamase with the third-generation antibiotic ceftriaxone by time-resolved serial femtosecond crystallography. The results reveal, in near atomic detail, antibiotic cleavage and inactivation from 30 ms to 2 s. Conclusions MISC is a versatile and generally applicable method to investigate reactions of biological macromolecules, some of which are of immense biological significance and might be, in addition, important targets for structure-based drug design. With megahertz X-ray pulse rates expected at the Linac Coherent Light Source II and the European X-ray free-electron laser, multiple, finely spaced time delays can be collected rapidly, allowing a comprehensive description of biomolecular reactions in terms of structure and kinetics from the same set of X-ray data.

Abstract Background Zika virus is an emerging pathogen of global importance. It has been responsible for recent outbreaks in the Americas and in the Pacific region. This study assessed five different mosquito species from the temperate climatic zone in Australia and included Aedes albopictus as a potentially invasive species. Methods Mosquitoes were orally challenged by membrane feeding with Zika virus strain of Cambodia 2010 origin, belonging to the Asian clade. Virus infection and dissemination were assessed by quantitative PCR on midgut and carcass after dissection. Transmission was assessed by determination of cytopathogenic effect of saliva (CPE) on Vero cells, followed by determination of 50% tissue culture infectious dose (TCID50) for CPE positive samples. Additionally, the presence of Wolbachia endosymbiont infection was assessed by qPCR and standard PCR. Results Culex mosquitoes were found unable to present Zika virus in saliva, as demonstrated by molecular as well as virological methods. Aedes aegypti, was used as a positive control for Zika infection and showed a high level of virus infection, dissemination and transmission. Local Aedes species, Ae. notoscriptus and, to a lesser degree, Ae. camptorhynchus were found to expel virus in their saliva and contained viral nucleic acid within the midgut. Molecular assessment identified low or no dissemination for these species, possibly due to low virus loads. Ae. albopictus from Torres Strait islands origin was shown as an efficient vector. Cx quinquefasciatus was shown to harbour Wolbachia endosymbionts at high prevalence, whilst no Wolbachia was found in Cx annulirostris. The Australian Ae. albopictus population was shown to harbour Wolbachia at high frequency. Conclusions The risk of local Aedes species triggering large Zika epidemics in the southern parts of Australia is low. The potentially invasive Ae. albopictus showed high prevalence of virus in the saliva and constitutes a potential threat if this mosquito species becomes established in mainland Australia. Complete risk analysis of Zika transmission in the temperate zone would require an assessment of the impact of temperature on Zika virus replication within local and invasive mosquito species.

Estrogen receptor alpha-positive (ERα+) luminal tumors are the most frequent subtype of breast cancer. Stat1−/− mice develop mammary tumors that closely recapitulate the biological characteristics of this cancer subtype. To identify transforming events that contribute to tumorigenesis, we performed whole genome sequencing of Stat1−/− primary mammary tumors and matched normal tissues. This investigation identified somatic truncating mutations affecting the prolactin receptor (PRLR) in all tumor and no normal samples. Targeted sequencing confirmed the presence of these mutations in precancerous lesions, indicating that this is an early event in tumorigenesis. Functional evaluation of these heterozygous mutations in Stat1−/− mouse embryonic fibroblasts showed that co-expression of truncated and wild-type PRLR led to aberrant STAT3 and STAT5 activation downstream of the receptor, cellular transformation in vitro, and tumor formation in vivo. In conclusion, truncating mutations of PRLR promote tumor growth in a model of human ERα+ breast cancer and warrant further investigation.

Mix-and-inject serial crystallography (MISC) is a technique designed to image enzyme catalyzed reactions in which small protein crystals are mixed with a substrate just prior to being probed by an X-ray pulse. This approach offers several advantages over flow cell studies. It provides (i) room temperature structures at near atomic resolution, (ii) time resolution ranging from microseconds to seconds, and (iii) convenient reaction initiation. It outruns radiation damage by using femtosecond X-ray pulses allowing damage and chemistry to be separated. Here, we demonstrate that MISC is feasible at an X-ray free electron laser by studying the reaction of M. tuberculosis ß-lactamase microcrystals with ceftriaxone antibiotic solution. Electron density maps of the apo-ß-lactamase and of the ceftriaxone bound form were obtained at 2.8 Å and 2.4 Å resolution, respectively. These results pave the way to study cyclic and non-cyclic reactions and represent a new field of time-resolved structural dynamics for numerous substrate-triggered biological reactions.

The phytoalexin trans-resveratrol, 5-[(1E)-2-(4-hydroxyphenyl)ethenyl]-1,3-benzenediol, is a well-known, potent antioxidant having a variety of possible biomedical applications. However, its adverse physicochemical properties (low stability, poor aqueous solubility) limit such applications and its inclusion in cyclodextrins (CDs) has potential for addressing these shortcomings. Here, various methods of the attempted synthesis of inclusion complexes between trans-resveratrol and three methylated cyclodextrins (permethylated α-CD, permethylated β-CD and 2,6-dimethylated β-CD) are described. Isolation of the corresponding crystalline 1:1 inclusion compounds enabled their full structure determination by X-ray analysis for the first time, revealing a variety of guest inclusion modes and unique supramolecular crystal packing motifs. The three crystalline inclusion complexes were also fully characterized by thermal analysis (hot stage microscopy, thermogravimetric analysis and differential scanning calorimetry). To complement the solid-state data, phase-solubility studies were conducted using a series of CDs (native and variously derivatised) to establish their effect on the aqueous solubility of trans-resveratrol and to estimate association constants for complex formation.

Our previous studies show reduced abundance of the β-subunit of mitochondrial H+-ATP synthase (β-F1-ATPase) in skeletal muscle of obese individuals. The β-F1-ATPase forms the catalytic core of the ATP synthase, and it is critical for ATP production in muscle. The mechanism(s) impairing β-F1-ATPase metabolism in obesity, however, are not completely understood. First, we studied total muscle protein synthesis and the translation efficiency of β-F1-ATPase in obese (BMI, 36±1 kg/m2) and lean (BMI, 22±1 kg/m2) subjects. Both total protein synthesis (0.044±0.006 vs 0.066±0.006%·h-1) and translation efficiency of β-F1-ATPase (0.0031±0.0007 vs 0.0073±0.0004) were lower in muscle from the obese subjects when compared to the lean controls (P

Although mosquitoes serve as vectors of many pathogens of public health importance, their response to viral infection is poorly understood. It also remains to be investigated whether viruses deploy some mechanism to be able to overcome this immune response. Here, we have used an RNA-Seq approach to identify differentially regulated genes in Culex quinquefasciatus cells following West Nile virus (WNV) infection, identifying 265 transcripts from various cellular pathways that were either upregulated or downregulated. Ubiquitin-proteasomal pathway genes, comprising 12% of total differentially regulated genes, were selected for further validation by real time RT-qPCR and functional analysis. It was found that treatment of infected cells with proteasomal inhibitor, MG-132, decreased WNV titers, indicating importance of this pathway during infection process. In infection models, the Culex ortholog of mammalian Cul4A/B (cullin RING ubiquitin ligase) was found to be upregulated in vitro as well as in vivo, especially in midguts of mosquitoes. Gene knockdown using dsRNA and overexpression studies indicated that Culex Cul4 acts as a pro-viral protein by degradation of CxSTAT via ubiquitin-proteasomal pathway. We also show that gene knockdown of Culex Cul4 leads to activation of the Jak-STAT pathway in mosquitoes leading to decrease viral replication in the body as well as saliva. Our results suggest a novel mechanism adopted by WNV to overcome mosquito immune response and increase viral replication.

BACKGROUND: Influenza A (flu) virus causes significant morbidity and mortality worldwide, and current vaccines require annual updating to protect against the rapidly arising antigenic variations due to antigenic shift and drift. In fact, current subunit or split flu vaccines rely exclusively on antibody responses for protection and do not induce cytotoxic T (Tc) cell responses, which are broadly cross-reactive between virus strains. We have previously reported that gamma-ray inactivated flu virus can induce cross-reactive Tc cell responses. METHODOLOGY/PRINCIPAL FINDING: Here, we report that intranasal administration of purified gamma-ray inactivated human influenza A virus preparations (gamma-Flu) effectively induces heterotypic and cross-protective immunity. A single intranasal administration of gamma-A/PR8[H1N1] protects mice against lethal H5N1 and other heterotypic infections. CONCLUSIONS/SIGNIFICANCE: Intranasal gamma-Flu represents a unique approach for a cross-protective vaccine against both seasonal as well as possible future pandemic influenza A virus infections.

Abstract Background Degradation of highly abundant stromal proteins plays an important role in the nitrogen economy of the plant during senescence. Lines of evidence supporting proteolysis within the chloroplast and outside the chloroplast have been reported. Two extra-plastidic degradation pathways, chlorophagy and Rubisco Containing Bodies, rely on cytoplasmic autophagy. Results In this work, levels of three stromal proteins (Rubisco large subunit, chloroplast glutamine synthetase and Rubisco activase) and one thylakoid protein (the major light harvesting complex protein of photosystem II) were measured during natural senescence in WT and in two autophagy T-DNA insertion mutants (atg5 and atg7). Thylakoid-localized protein decreased similarly in all genotypes, but stromal protein degradation was incomplete in the two atg mutants. In addition, degradation of two stromal proteins was observed in chloroplasts isolated from mid-senescence leaves. Conclusions These data suggest that autophagy does contribute to the complete proteolysis of stromal proteins, but does not play a major degenerative role. In addition, support for in organello degradation is provided.

Abstract Background The trafficking of the adenomatous polyposis coli (APC) tumour suppressor protein in mammalian cells is a perennially controversial topic. Immunostaining evidence for an actin-associated APC localisation at intercellular junctions has been previously presented, though live imaging of mammalian junctional APC has not been documented. Results Using live imaging of transfected COS-7 cells we observed intercellular junction-associated pools of GFP-APC in addition to previously documented microtubule-associated GFP-APC and a variety of minor localisations. Although both microtubule and junction-associated populations could co-exist within individual cells, they differed in their subcellular location, dynamic behaviour and sensitivity to cytoskeletal poisons. GFP-APC deletion mutant analysis indicated that a protein truncated immediately after the APC armadillo repeat domain retained the ability to localise to adhesive membranes in transfected cells. Supporting this, we also observed junctional APC immunostaining in cultures of human colorectal cancer cell line that express truncated forms of APC. Conclusion Our data indicate that APC can be found in two spatially separate populations at the cell periphery and these populations can co-exist in the same cell. The first localisation is highly dynamic and associated with microtubules near free edges and in cell vertices, while the second is comparatively static and is closely associated with actin at sites of cell-cell contact. Our imaging confirms that human GFP-APC possesses many of the localisations and behaviours previously seen by live imaging of Xenopus GFP-APC. However, we report the novel finding that GFP-APC puncta can remain associated with the ends of shrinking microtubules. Deletion analysis indicated that the N-terminal region of the APC protein mediated its junctional localisation, consistent with our observation that truncated APC proteins in colon cancer cell lines are still capable of localising to the cell cortex. This may have implications for the development of colorectal cancer.

Abstract Background EB1 is a microtubule tip-associated protein that interacts with the APC tumour suppressor protein and the p150glued subunit of dynactin. We previously reported that an EB1 deletion mutant that retains both of these interactions but does not directly associate with microtubules (EB1-ΔN2-GFP) spontaneously formed perinuclear aggregates when expressed in COS-7 cells. Results In the present study live imaging indicated that EB1-ΔN2-GFP aggregates underwent dynamic microtubule-dependent changes in morphology and appeared to be internally cohesive. EB1-ΔN2-GFP aggregates were phase-dense structures that displayed microtubule-dependent accumulation around the centrosome, were immunoreactive for both the 20s subunit of the proteasome and ubiquitin, and induced the collapse of the vimentin cytoskeleton. Fractionation studies revealed that a proportion of EB1-ΔN2-GFP was detergent-insoluble and ubiquitylated, indicating that EB1-ΔN2-GFP aggregates are aggresomes. Immunostaining also revealed that APC and p150glued were present in EB1-ΔN2-GFP aggregates, whereas EB3 was not. Furthermore, evidence for p150glued degradation was found in the insoluble fraction of EB1-ΔN2-GFP transfected cultures. Conclusion Our data indicate that aggresomes can be internally cohesive and may not represent a simple "aggregate of aggregates" assembled around the centrosome. Our observations also indicate that a partially misfolded protein may retain the ability to interact with its normal physiological ligands, leading to their co-assembly into aggresomes. This supports the idea that the trapping and degradation of co-aggregated proteins might contribute to human pathologies characterised by aggresome formation.

Background: The endangered Formosan black bear (Ursus thibetanus formosanus) is the largest native carnivorous mammal in Taiwan. Diseases, poor management, illegal hunting, and habitat destruction are serious threats to the survival of bear populations. However, studies on the impact of diseases on bear populations are limited. Therefore, this study aimed to establish a database of the hematological and plasma profiles of free-ranging Formosan black bears and investigate the occurrence of ectoparasites, blood parasites, and vector-borne pathogens., Methods: Formosan black bears were captured in Yushan National Park (YNP) and Daxueshan Forest Recreation Area (DSY) in Taiwan. Blood samples were collected from each bear for hematological analysis and plasma biochemistry using a hematology analyzer. Parasites and pathogens were detected using a thin blood smear with Wright-Giemsa staining and polymerase chain reaction (PCR) assay. Additionally, macroscopic ectoparasites were collected from bears to detect blood parasites and other pathogens. Moreover, the relationships between the bear variables (sex, age, and occurrence of parasites or pathogens), ectoparasites, and infectious agents were also analyzed., Results: In all, 21 wild bears (14 in YNP and 7 in DSY) were captured and released during the satellite tracking studies. Hematological analysis and plasma biochemistry indicated significant differences in white blood cells (WBC), segments, creatine kinase (CK), and lactate dehydrogenase (LDH) levels between foot snare and culvert-captured bears. Additionally, there were significant differences in total plasma protein (TPP), creatinine, Ca 2+ , Mg 2+ , and K + levels between male and female bears. Moreover, pathogen-infected bears had significantly higher erythrocyte sedimentation rate (ESR; 30 min and 1 h) and globulin levels than uninfected bears. In total, 240 ticks were collected from 13 bears, among which eight adult tick species were identified, including Haemaphysalis flava, Haemaphysalis hystricis, Amblyomma testudinarium, Ixodes ovatus, Dermacentor taiwanensis, Haemaphysalis longicornis, Ixodes acutitarsus, Amblyomma javanense, and nymphs belonging to Haemaphysalis spp. PCR revealed that 13 (61.90%) and 8 (38.10%) bears harbored Hepatozoon ursi and Babesia DNA, respectively. Among the ticks examined, 157 (65.41%) and 128 (53.33%) samples were positive for H. ursi and Babesia, respectively., Conclusions: To the best of our knowledge, this is the first study to establish a database of the hematological and plasma profiles of wild Formosan black bears and investigate ectoparasite infestation and Hepatozoon and Babesia spp., Infection: In conclusion, these findings may serve as a reference for monitoring the health and population of locally endangered bears., (© 2024. The Author(s).)

Despite the capability of extracellular vesicles (EVs) derived from Gram-negative and Gram-positive bacteria to induce potent anti-tumour responses, large-scale production of bacterial EVs remains as a hurdle for their development as novel cancer immunotherapeutic agents. Here, we developed manufacturing processes for mass production of Escherichia coli EVs, namely, outer membrane vesicles (OMVs). By combining metal precipitation and size-exclusion chromatography, we isolated 357 mg in total protein amount of E. coli OMVs, which was equivalent to 3.93 × 10 15 particles (1.10 × 10 10 particles/μg in total protein amounts of OMVs) from 160 L of the conditioned medium. We show that these mass-produced E. coli OMVs led to complete remission of two mouse syngeneic tumour models. Further analysis of tumour microenvironment in neoantigen-expressing tumour models revealed that E. coli OMV treatment causes increased infiltration and activation of CD8 + T cells, especially those of cancer antigen-specific CD8 + T cells with high expression of TCF-1 and PD-1. Furthermore, E. coli OMVs showed synergistic anti-tumour activity with anti-PD-1 antibody immunotherapy, inducing substantial tumour growth inhibition and infiltration of activated cancer antigen-specific stem-like CD8 + T cells into the tumour microenvironment. These data highlight the potent anti-tumour activities of mass-produced E. coli OMVs as a novel candidate for developing next-generation cancer immunotherapeutic agents., (© 2023 The Authors. Journal of Extracellular Vesicles published by Wiley Periodicals, LLC on behalf of the International Society for Extracellular Vesicles.)

Multi-cancer early detection remains a key challenge in cell-free DNA (cfDNA)-based liquid biopsy. Here, we perform cfDNA whole-genome sequencing to generate two test datasets covering 2125 patient samples of 9 cancer types and 1241 normal control samples, and also a reference dataset for background variant filtering based on 20,529 low-depth healthy samples. An external cfDNA dataset consisting of 208 cancer and 214 normal control samples is used for additional evaluation. Accuracy for cancer detection and tissue-of-origin localization is achieved using our algorithm, which incorporates cancer type-specific profiles of mutation distribution and chromatin organization in tumor tissues as model references. Our integrative model detects early-stage cancers, including those of pancreatic origin, with high sensitivity that is comparable to that of late-stage detection. Model interpretation reveals the contribution of cancer type-specific genomic and epigenomic features. Our methodologies may lay the groundwork for accurate cfDNA-based cancer diagnosis, especially at early stages., (© 2023. The Author(s).)

With advances in next-generation sequencing technology, non-invasive prenatal testing (NIPT) has been widely implemented to detect fetal aneuploidies, including trisomy 21, 18, and 13 (T21, T18, and T13). Most NIPT methods use cell-free DNA (cfDNA) fragment count (FC) in maternal blood. In this study, we developed a novel NIPT method using cfDNA fragment distance (FD) and convolutional neural network-based artificial intelligence algorithm (aiD-NIPT). Four types of aiD-NIPT algorithm (mean, median, interquartile range, and its ensemble) were developed using 2,215 samples. In an analysis of 17,678 clinical samples, all algorithms showed >99.40% accuracy for T21/T18/T13, and the ensemble algorithm showed the best performance (sensitivity: 99.07%, positive predictive value (PPV): 88.43%); the FC-based conventional Z-score and normalized chromosomal value showed 98.15% sensitivity, with 40.77% and 36.81% PPV, respectively. In conclusion, FD-based aiD-NIPT was successfully developed, and it showed better performance than FC-based NIPT methods., Competing Interests: Authors JL, S-ML, JMA, T-RL, WK, E-HC, and C-SK were employed by GC Genome., (Copyright © 2022 Lee, Lee, Ahn, Lee, Kim, Cho and Ki.)

Genetic mutations can cause life-threatening diseases such as cancers and sickle cell anemia. Gene detection is thus of importance for disease-risk prediction or early diagnosis and treatment. Apart from genetic defects, gene detection techniques can also be applied to gene-related diseases with high risk to human health such as human papillomavirus (HPV) infection. HPV infection has been strongly linked to cervical cancer. To achieve a high-throughput HPV gene detection platform, the flow-through hybridization system appears to be one of the commercialized diagnostic techniques for this purpose. The flow-through hybridization technique is based on a vacuum-guided flow of DNA fragments which is continuously directed toward the oligoprobes that are immobilized on the testing membrane. However, the conventional colorimetric method and signal read-out approach suffers a problem of low sensitivity. On the contrary, fluorescence approaches allow more sensitive detection and broad sensing ranges. In this work, a fluorescent dye HCAP, which possesses aggregation-induced emission (AIE) properties and is responsive to alkaline phosphatase, was developed and applied to the flow-through hybridization platform to achieve HPV genome diagnosis of clinical samples. Also, an automatic membrane reader was constructed based on the AIE-based diagnosis platform which can identify the diagnostic result of patient DNA with a total concordance rate of 100% in the clinical trial.

Background: Statins are the most popular agents for the primary and secondary prevention of cardiovascular disease; however, the pharmacokinetic parameters and associated genetic factors in the Korean population have not been fully elucidated. This study explored the pharmacokinetic properties of atorvastatin and the association between genetic variations and atorvastatin pharmacokinetics in healthy Korean subjects. Methods: Atorvastatin (80 mg) was administered to 35 healthy Korean volunteers. Plasma levels of atorvastatin and its metabolites were measured sequentially using liquid chromatography-tandem mass spectrometry from 0 to 24 h after atorvastatin administration. Customized next-generation sequencing analysis was performed covering all coding exons of 15 genes, as well as 46 single-nucleotide variants in 29 genes related to statin pharmacokinetics. Results: The mean area under the concentration-time (AUC) and C max (maximum peak concentration) were 269.0 ng/ml∙h and 84.3 ng/ml, respectively, which were approximately two times higher than those reported in Caucasians. Genetic analysis revealed that eight genetic variants in ABCB1, ABCG2, APOA5, CETP , and CYP7A1 contributed to the AUC of atorvastatin. The atorvastatin AUC 0-24 h prediction model was developed based on age and eight genetic variants using multivariate linear regression (adjusted R 2 = 0.878, p < 0.0001). Conclusion: This study shows that the pharmacokinetic properties of atorvastatin in Koreans are different from those in Caucasians and that atorvastatin AUC 0-24 h could be predicted based on age and eight genetic variants of ABCB1, ABCG2, APOA5, CETP, and CYP7A1 ., Competing Interests: Authors TL, T-RL, E-HC, and C-SK were employed by the company GC Genome. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Kim, Seo, Yun, Kim, Kim, Lee, Lee, Lee, Cho and Ki.)

Telomerase is essential for the immortality characteristics of most cancers. Telomerase-specific inhibitors should render cancer cells to replicative senescence without acute cytotoxicity. Perylene-based G-quadruplex (G4) ligands are widely studied as telomerase inhibitors. Most reported perylene-based G4 ligands are perylene diimides (PDIs), which often suffer from self-aggregation in aqueous solutions. Previously, we found that PM2, a perylene monoimide (PMI), exhibited better solubility, G4 binding affinity, and telomerase inhibition than PIPER, the prototypic PDI. However, the acute cytotoxicity of PM2 was about 20-30 times more than PIPER in cancer cells. In this report, we replaced the piperazine side chain of PM2 with ethylenediamine to yield PM3 and replaced the N , N -diethylethylenediamine side chain of PM2 with the 1-(2-aminoethyl) piperidine to yield PM5. We found that asymmetric PMIs with two basic side chains (PM2, PM3, and PM5) performed better than PIPER (the prototypic PDI), in terms of hydrosolubility, G4 binding, in vitro telomerase inhibition, and suppression of human telomerase reverse transcriptase ( hTERT ) expression and telomerase activity in A549 cells. However, PM5 was 7-10 times less toxic than PM2 and PM3 in three cancer cell lines. We conclude that replacing the N , N -diethylethylenediamine side chain with the 2-aminoethylpiperidine on PMIs reduces the cytotoxicity in cancer cells without impacting G4 binding and telomerase inhibition. This study paves the way for synthesizing new PMIs with drug-like properties for selective telomerase inhibition., Competing Interests: The authors declare no competing financial interest., (© 2022 The Authors. Published by American Chemical Society.)

Due to their good magnetic properties, excellent biocompatibility, and low price, magnetic iron oxide nanoparticles (IONPs) are the most commonly used magnetic nanomaterials and have been extensively explored in biomedical applications. Although magnetic IONPs can be used for a variety of applications in biomedicine, most practical applications require IONP-based platforms that can perform several tasks in parallel. Thus, appropriate engineering and integration of magnetic IONPs with different classes of organic and inorganic materials can produce multifunctional nanoplatforms that can perform several functions simultaneously, allowing their application in a broad spectrum of biomedical fields. This review article summarizes the fabrication of current composite nanoplatforms based on integration of magnetic IONPs with organic dyes, biomolecules (e.g., lipids, DNAs, aptamers, and antibodies), quantum dots, noble metal NPs, and stimuli-responsive polymers. We also highlight the recent technological advances achieved from such integrated multifunctional platforms and their potential use in biomedical applications, including dual-mode imaging for biomolecule detection, targeted drug delivery, photodynamic therapy, chemotherapy, and magnetic hyperthermia therapy.

Magnetite (Fe 3 O 4 ) nanoparticles (NPs) are attractive nanomaterials in the field of material science, chemistry, and physics because of their valuable properties, such as soft ferromagnetism, half-metallicity, and biocompatibility. Various structures of Fe 3 O 4 NPs with different sizes, geometries, and nanoarchitectures have been synthesized, and the related properties have been studied with targets in multiple fields of applications, including biomedical devices, electronic devices, environmental solutions, and energy applications. Tailoring the sizes, geometries, magnetic properties, and functionalities is an important task that determines the performance of Fe 3 O 4 NPs in many applications. Therefore, this review focuses on the crucial aspects of Fe 3 O 4 NPs, including structures, synthesis, magnetic properties, and strategies for functionalization, which jointly determine the application performance of various Fe 3 O 4 NP-based systems. We first summarize the recent advances in the synthesis of magnetite NPs with different sizes, morphologies, and magnetic properties. We also highlight the importance of synthetic factors in controlling the structures and properties of NPs, such as the uniformity of sizes, morphology, surfaces, and magnetic properties. Moreover, emerging applications using Fe 3 O 4 NPs and their functionalized nanostructures are also highlighted with a focus on applications in biomedical technologies, biosensing, environmental remedies for water treatment, and energy storage and conversion devices., Competing Interests: Conflicts of Interest: The authors have no competing financial interest to declare.

Genomic instability of circulating tumor DNA (ctDNA) as a prognostic biomarker has not been evaluated in pancreatic cancer. We investigated the role of the genomic instability index of ctDNA in pancreatic ductal adenocarcinoma (PDAC). We prospectively enrolled 315 patients newly diagnosed with resectable ( n = 110), locally advanced ( n = 78), and metastatic ( n = 127) PDAC from March 2015 through January 2020. Low-depth whole-genome cell-free DNA sequencing identified genome-wide copy number alterations using instability score (I-score) to reflect genome-wide instability. Plasma cell-free and matched tumor tissue DNA from 15 patients with resectable pancreatic cancer was sequenced to assess the concordance of chromosomal copy number alteration profiles. Associations of I-score with clinical factors or survival were assessed. Seventy-six patients had high genomic instability with I-score > 7.3 in pre-treatment ctDNA; proportions of high I-score were 5.5%, 5.1%, and 52% in resectable, locally advanced, and metastatic stages, respectively. Correlation coefficients between Z-scores of plasma and tissue DNA at segment resolution were high (r 2 = 0.82). Univariable analysis showed the association of I-score with progression-free survival in each stage. Multivariable analyses demonstrated that clinical stage-adjusted I-scores were significant factors for progression-free and overall survival. In these patients, ctDNA genomic I-scores provided prognostic information relevant to progression-free survival in each clinical stage.

Traumatic brain injury (TBI) is a major public health challenge that is also the third leading cause of death worldwide. It is also the leading cause of long-term disability in children and young adults worldwide. Despite a large body of research using predominantly in vivo and in vitro rodent models of brain injury, there is no medication that can reduce brain damage or promote brain repair mainly due to our lack of understanding in the mechanisms and pathophysiology of the TBI. The aim of this review is to examine in vitro TBI studies conducted from 2008-2018 to better understand the TBI in vitro model available in the literature. Specifically, our focus was to perform a detailed analysis of the in vitro experimental protocols used and their subsequent biological findings. Our review showed that the uniaxial stretch is the most frequently used way of load application, accounting for more than two-thirds of the studies reviewed. The rate and magnitude of the loading were varied significantly from study to study but can generally be categorized into mild, moderate, and severe injuries. The in vitro studies reviewed here examined key processes in TBI pathophysiology such as membrane disruptions leading to ionic dysregulation, inflammation, and the subsequent damages to the microtubules and axons, as well as cell death. Overall, the studies examined in this review contributed to the betterment of our understanding of TBI as a disease process. Yet, our review also revealed the areas where more work needs to be done such as: 1) diversification of load application methods that will include complex loading that mimics in vivo head impacts; 2) more widespread use of human brain cells, especially patient-matched human cells in the experimental set-up; and 3) need for building a more high-throughput system to be able to discover effective therapeutic targets for TBI.

The Sewol Ferry Disaster which took place in 16th of April, 2014, was a national level disaster in South Korea that caused severe social distress nation-wide. No research at the domestic level thus far has examined the influence of the disaster on social stress through a sentiment analysis of social media data. Data extracted from YouTube, Twitter, and Facebook were used in this study. The population was users who were randomly selected from the aforementioned social media platforms who had posted texts related to the disaster from April 2014 to March 2015. ANOVA was used for statistical comparison between negative, neutral, and positive sentiments under a 95% confidence level. For NLP-based data mining results, bar graph and word cloud analysis as well as analyses of phrases, entities, and queries were implemented. Research results showed a significantly negative sentiment on all social media platforms. This was mainly related to fundamental agents such as ex-president Park and her related political parties and politicians. YouTube, Twitter, and Facebook results showed negative sentiment in phrases (63.5, 69.4, and 58.9%, respectively), entity (81.1, 69.9, and 76.0%, respectively), and query topic (75.0, 85.4, and 75.0%, respectively). All results were statistically significant ( p < 0.001). This research provides scientific evidence of the negative psychological impact of the disaster on the Korean population. This study is significant because it is the first research to conduct sentiment analysis of data extracted from the three largest existing social media platforms regarding the issue of the disaster., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2020 Lee, Lee, Lee, Jang and Kim.)

Background: Early detection of childhood developmental delays is very important for the treatment of disabilities., Objective: To investigate the possibility of detecting childhood developmental delays leading to disabilities before clinical registration by analyzing big data from a health insurance database., Methods: In this study, the data from children, individuals aged up to 13 years (n=2412), from the Sample Cohort 2.0 DB of the Korea National Health Insurance Service were organized by age range. Using 6 categories (having no disability, having a physical disability, having a brain lesion, having a visual impairment, having a hearing impairment, and having other conditions), features were selected in the order of importance with a tree-based model. We used multiple classification algorithms to find the best model for each age range. The earliest age range with clinically significant performance showed the age at which conditions can be detected early., Results: The disability detection model showed that it was possible to detect disabilities with significant accuracy even at the age of 4 years, about a year earlier than the mean diagnostic age of 4.99 years., Conclusions: Using big data analysis, we discovered the possibility of detecting disabilities earlier than clinical diagnoses, which would allow us to take appropriate action to prevent disabilities., (©Seung-Hyun Jeong, Tae Rim Lee, Jung Bae Kang, Mun-Taek Choi. Originally published in JMIR Medical Informatics (http://medinform.jmir.org), 23.11.2020.)

Perylene diimide (PDI) derivatives have been studied as G-quadruplex ligands that suppress telomerase activity by facilitating G-quadruplex formation of telomeric DNA and the hTERT promoter. PIPER, the prototypical PDI, reduces telomerase activity in lung and prostate cancer cells, leading to telomere shortening and cellular senescence of these cells. However, PIPER suffers from poor hydrosolubility and the propensity to aggregate at neutral pH. In this report, we synthesized a new asymmetric PDI, aPDI-PHis, which maintains one N -ethyl piperidine side chain of PIPER and has histidine as another side chain. The results show that aPDI-PHis is superior to its symmetric counterparts, PIPER and PDI-His, in terms of hydrosolubility, G-quadruplex binding, cellular uptake, and telomerase inhibition in prostate cancer cells. These results suggest that one N -ethyl piperidine side chain of PDI is sufficient for G-quadruplex binding, while another side chain can be tuned to elicit desirable properties. These findings might lead to better PDIs for use as anticancer drugs., Competing Interests: The authors declare no competing financial interest., (© 2020 American Chemical Society.)

This article details the preparation of hollow gold-silver nanoshells (GS-NSs) coated with tunably thin silica shells for use in plasmon-enhanced photocatalytic applications. Hollow GS-NSs were synthesized via the galvanic replacement of silver nanoparticles. The localized surface plasmon resonance (LSPR) peaks of the GS-NSs were tuned over the range of visible light to near-infrared (NIR) wavelengths by adjusting the ratio of silver nanoparticles to gold salt solution to obtain three distinct types of GS-NSs with LSPR peaks centered near 500, 700, and 900 nm. Varying concentrations of (3-aminopropyl)trimethoxysilane and sodium silicate solution afforded silica shell coatings of controllable thicknesses on the GS-NS cores. For each type of GS-NS, scanning electron microscopy (SEM) and transmission electron microscopy (TEM) images verified our ability to grow thin silica shells having three different thicknesses of silica shell (~2, ~10, and ~15 nm) on the GS-NS cores. Additionally, energy-dispersive X-ray (EDX) spectra confirmed the successful coating of the GS-NSs with SiO 2 shells having controlled thicknesses. Extinction spectra of the as-prepared nanoparticles indicated that the silica shell has a minimal effect on the LSPR peak of the gold-silver nanoshells.

Core@shell metal nanoparticles have emerged as promising photocatalysts because of their strong and tunable plasmonic properties; however, marked improvements in photocatalytic efficiency are needed if these materials are to be widely used in practical applications. Accordingly, the design of new and functional light-responsive nanostructures remains a central focus of nanomaterial research. To this end, we report the synthesis of nanorattles comprising hollow gold-silver nanoshells encapsulated within vacuous tin oxide shells of adjustable thicknesses (∼10 and ∼30 nm for the two examples prepared in this initial report). These composite nanorattles exhibited broad tunable optical extinctions ranging from ultraviolet to near-infrared spectral regions (i.e., 300-745 nm). Zeta potential measurements showed a large negative surface charge of approximately -35 mV, which afforded colloidal stability to the nanorattles in aqueous solution. We also characterized the nanorattles structurally and compositionally using scanning electron microscopy, transmission electron microscopy, and energy-dispersive X-ray spectroscopy. Futhermore, finite-difference time-domain simulation and photoluminescence properties of the composited nanoparticles were investigated. Collectively, these studies indicate that our tin oxide-coated hollow gold-silver nanorattles are promising candidates for use in solar-driven applications., Competing Interests: The authors declare no competing financial interest., (Copyright © 2020 American Chemical Society.)

New ether-substituted poly(1,4-phenylene vinylene) (PPV) derivatives were synthesized via Horner-Emmons coupling. The structures of the monomers and the resultant oligomers were confirmed by 1 H and 13 C NMR spectroscopies. The molecular weights of the oligomers were characterized by gel permeation chromatography, giving the number-average and weight-average molecular weights and the corresponding polydispersity indices. Measurements of UV-vis absorption and fluorescence were used to characterize the optical properties of the oligomers. Estimation of the highest occupied molecular orbital and lowest unoccupied molecular orbital energy levels and other electrochemical characteristics of the oligomers were investigated by cyclic voltammetry. Dialkyl and dialkoxy PPV oligomers were also prepared and characterized following the same instrumental methods used for the ether-substituted oligomers, providing a known reference system to judge the performance of the new conjugated oligomers. Devices were fabricated to analyze the electroluminescent characteristics of the oligomers in organic light-emitting diodes., Competing Interests: The authors declare no competing financial interest., (Copyright © 2019 American Chemical Society.)

This paper describes the preparation and characterization of a new type of core-shell nanoparticle in which the structure consists of a hydrogel core encapsulated within a porous silver shell. The thermo-responsive hydrogel cores were prepared by surfactant-free emulsion polymerization of a selected mixture of N -isopropylacrylamide (NIPAM) and acrylic acid (AAc). The hydrogel cores were then encased within either a porous or complete silver shell for which the localized surface plasmon resonance (LSPR) extends from visible to near-infrared (NIR) wavelengths (i.e., λ max varies from 550 to 1050 nm, depending on the porosity), allowing for reversible contraction and swelling of the hydrogel via photothermal heating of the surrounding silver shell. Given that NIR light can pass through tissue, and the silver shell is porous, this system can serve as a platform for the smart delivery of payloads stored within the hydrogel core. The morphology and composition of the composite nanoparticles were characterized by SEM, TEM, and FTIR, respectively. UV-vis spectroscopy was used to characterize the optical properties., (Copyright © 2019, Bryan et al.; licensee Beilstein-Institut.)

A "smart" core@shell composite nanoparticle (NP) having dual-response mechanisms (i.e., temperature and light) was synthesized, and its efficacy in the loading and release of small molecules was explored. These core@shell NPs are composed of an optically active gold nanoshell (GNS) core and a mesoporous ( m -) silica layer ( m -SiO 2 ). The GNS@ m -SiO 2 nanoparticles are further encapsulated within a thermo-responsive poly(N-isopropylacrylamide-co-acrylic acid) hydrogel (PNIPAM-co-AA). The multi-responsive composite NPs were designed to create thermally and optically modulated drug-delivery vehicles with a m -SiO 2 layer providing additional non-collapsible space for drug storage. The influence of the m -SiO 2 layer on the efficacy of loading and release of methylene blue, which serves as a model for a small-molecule therapeutic drug, was evaluated. The "smart" core@shell composite NPs having a m -SiO 2 layer demonstrated an improved capacity to load and release small molecules compared to the corresponding NPs with no m -SiO 2 shell. Additionally, an efficient response by the composite NPs was successfully induced by the thermal energy generated from the gold nanoshell core upon exposure to near infrared (NIR) stimulation.

Prostate cancer is the second most common cancer among men worldwide, and it is ranked first in the United States and Europe. Since prostate cancer is slow-growing, active surveillance for low-risk cancer has been increasingly supported by various guidelines. Most prostate cancers reactivate telomerase to circumvent the replicative senescence caused by the end replication problem; therefore, telomerase inhibition is potentially useful for the suppression of prostate cancer progression during this active surveillance or for the prevention of cancer recurrence after conventional therapies. In this study, we demonstrated that the perylene derivatives, PM2 and PIPER, could suppress human telomerase reverse transcriptase (hTERT) expression and telomerase activity in the short-term treatment of androgen-dependent prostate cancer cell line LNCaP and the androgen-independent prostate cancer cell line PC3 prostate cancer cells. Long-term treatment with subcytotoxic doses of these compounds in both prostate cancer cells showed telomere shortening and a significant increase in senescent cells. Although the acute cytotoxicity of PM2 was about 30 times higher than that of PIPER in both prostate cancer cells, the cellular uptake of both compounds was comparable as determined by flow cytometry and fluorescent microscopy.

The authors wish to make the following correction to this paper [...].

Dehydroabietic acid (DAA) is a naturally occurring diterpene resin acid derived from coniferous plants such as Pinus and Picea . Various bioactive effects of DAA have been studied including antibacterial, antifungal, and anticancer activities. However, the anti-inflammatory mechanism of DAA remains unclear. We evaluated the anti-inflammatory effect of DAA in macrophage cell lines. Dehydroabietic acid clearly reduced nitric oxide (NO) production and inflammatory gene expression decreased according to RT-PCR results. Dehydroabietic acid displayed anti-inflammatory activity at the transcriptional level in results from NF-κB- or AP-1-mediated luciferase assays. To identify the DAA target protein, we investigated NF-κB and AP-1 pathways by Western blotting analysis. Dehydroabietic acid suppressed the activity of proto-oncogene tyrosine protein kinase (Src) and spleen tyrosine kinase (Syk) in the NF-κB cascade and transforming growth factor beta-activated kinase 1 (TAK1) in the AP-1 cascade. Using overexpression strategies, we confirmed that DAA targeted these kinases. Our findings demonstrate the anti-inflammatory effects and molecular mechanism of DAA. This suggests that DAA has potential as a drug or supplement to ameliorate inflammation., Competing Interests: The authors declare no conflict of interest

Exposure to fine particulate matter (PM) with diameter <2.5 µm (PM2.5) causes epithelium injury and endothelial dysfunction. Primary cilia are sensory organelles that transmit extracellular signals into intracellular biochemical responses and have roles in physiology. To date, there have been no studies investigating whether PM2.5 affects primary cilia in skin. We addressed this in the present study using normal human epidermal keratinocytes (NHEKs) and retinal pigment epithelium (RPE) cells. We found that formation of primary cilium is increased in differentiated NHEKs. However, treatment with PM2.5 blocked increased ciliogenesis in NHEKs and RPE cells. Furthermore, PM2.5 transcriptionally upregulated small proline rich protein 3 (SPRR3) expression by activating c-Jun, and ectopic expression of SPRR3 inhibits suppressed the ciliogenesis. Accordingly, treatment with c-Jun activator (anisomycin) induced SPRR3 expression, whereas the inhibitor (SP600125) recovered the ciliated cells and cilium length in PM2.5-treated cells. Moreover, c-Jun inhibitor suppressed upregulation of SPRR3 in PM2.5-treated cells. Taken together, our finding suggested that PM2.5 inhibits ciliogenesis by increasing SPRR3 expression via c-Jun activation in RPE cells and keratinocytes.

Mannosylerythritol lipids (MELs) are glycolipids and have several pharmacological efficacies. MELs also show skin-moisturizing efficacy through a yet-unknown underlying mechanism. Aquaporin-3 (AQP3) is a membrane protein that contributes to the water homeostasis of the epidermis, and decreased AQP3 expression following ultraviolet (UV)-irradiation of the skin is associated with reduced skin moisture. No previous study has examined whether the skin-moisturizing effect of MELs might act through the modulation of AQP3 expression. Here, we report for the first time that MELs ameliorate the UVA-induced downregulation of AQP3 in cultured human epidermal keratinocytes (HaCaT keratinocytes). Our results revealed that UVA irradiation decreases AQP3 expression at the protein and messenger RNA (mRNA) levels, but that MEL treatment significantly ameliorated these effects. Our mitogen-activated protein kinase inhibitor analysis revealed that phosphorylation of c-Jun N-terminal kinase (JNK), but not extracellular signal-regulated kinase or p38, mediates UVA-induced AQP3 downregulation, and that MEL treatment significantly suppressed the UVA-induced phosphorylation of JNK. To explore a possible mechanism, we tested whether MELs could regulate the expression of peroxidase proliferator-activated receptor gamma (PPAR-γ), which acts as a potent transcription factor for AQP3 expression. Interestingly, UVA irradiation significantly inhibited the mRNA expression of PPAR-γ in HaCaT keratinocytes, whereas a JNK inhibitor and MELs significantly rescued this effect. Taken together, these findings suggest that MELs ameliorate UVA-induced AQP3 downregulation in HaCaT keratinocytes by suppressing JNK activation to block the decrease of PPAR-γ. Collectively, our findings suggest that MELs can be used as a potential ingredient that modulates AQP3 expression to improve skin moisturization following UVA irradiation-induced damage., Competing Interests: CONFLICTS OF INTEREST: The authors declare no conflicts of interest.

Late epidermal differentiation is a key step of skin barrier formation; however, the specific genetic factors that distinguish late differentiation from early differentiation remain unknown. Here, we demonstrated that EGR3 is highly expressed in the stratum granulosum, and that it contributes to late epidermal differentiation. However, its expression is lost under poorly differentiated conditions, such as parakeratosis-lesional skin. EGR3 mediated the regulation of genes located in the epidermal differentiation complex through activation of enhancers and induction of enhancer RNAs. We further identified 20 targets of EGR3 specific for late differentiation. Additionally, we discovered that EGR3- and EGR3-related genes exhibited high tissue specificity on the skin. Through weighted gene co-expression analysis, EGR3 was found to be related to the keratinocyte differentiation-related module as an important part of the skin-specific genetic network. These findings shed light on the transcriptional regulation of late epidermal differentiation, highlighting candidate targets for diseases related to disrupted differentiation., (Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.)