N6-methyladenosine demethylase fat mass and obesity-associated protein suppresses hyperglycemia-induced endothelial cell injury by inhibiting reactive oxygen species formation via autophagy promotion.

INTRODUCTION: Hyperglycemia-induced endothelial cell injury is one of the main causes of diabetic vasculopathy. Fat mass and obesity-associated protein (FTO) was the first RNA N6-methyladenosine (m6A) demethylase identified; it participates in the pathogenesis of diabetes. However, the role of FTO in hyperglycemia-induced vascular endothelial cell injury remains unclear. MATERIALS AND METHODS: The effects of FTO on cellular m6A, autophagy, oxidative stress, proliferation, and cytotoxicity were explored in human umbilical vein endothelial cells (HUVECs) treated with high glucose (33.3 mmol/mL) after overexpression or pharmacological inhibition of FTO. MeRIP-qPCR and RNA stability assays were used to explore the molecular mechanisms by which FTO regulates autophagy. RESULTS: High glucose treatment increased m6A levels and reduced FTO protein expression in HUVECs. Wild-type overexpression of FTO markedly inhibited reactive oxygen species generation by promoting autophagy, increasing endothelial cell proliferation, and decreasing the cytotoxicity of high glucose concentrations. The pharmacological inhibition of FTO showed the opposite results. Mechanistically, we identified Unc-51-like kinase 1 (ULK1), a gene responsible for autophagosome formation, as a downstream target of FTO-mediated m6A modification. FTO overexpression demethylated ULK1 mRNA and inhibited its degradation in an m6A-YTHDF2-dependent manner, leading to autophagy activation. CONCLUSIONS: Our study demonstrates the functional importance of FTO-mediated m6A modification in alleviating endothelial cell injury under high glucose conditions and indicates that FTO may be a novel therapeutic target for diabetic vascular complications.

The single cell immunogenomic landscape after neoadjuvant immunotherapy combined chemotherapy in esophageal squamous cell carcinoma.

Neoadjuvant immunotherapy represents promising strategy in the treatment of esophageal squamous cell carcinoma (ESCC). However, the mechanisms underlying its impact on treatment sensitivity or resistance remain a subject of controversy. In this study, we conducted single-cell RNA and T/B cell receptor (scTCR/scBCR) sequencing of CD45+ immune cells on samples from 10 patients who received neoadjuvant immunotherapy and chemotherapy. We also validated our findings using multiplexed immunofluorescence and analyzed bulk RNA-seq from other cohorts in public database. By integrating analysis of 87357 CD45+ cells, we found GZMK + effector memory T cells (Tem) were relatively enriched and CXCL13+ exhausted T cells (Tex) and regulator T cells (Treg) decreased among responders, indicating a persistent anti-tumor memory process. Additionally, the enhanced presence of BCR expansion and somatic hypermutation process within TNFRSF13B + memory B cells (Bmem) suggested their roles in antigen presentation. This was further corroborated by the evidence of the T-B co-stimulation pattern and CXCL13-CXCR5 axis. The complexity of myeloid cell heterogeneity was also particularly pronounced. The elevated expression of S100A7 in ESCC, as detected by bulk RNA-seq, was associated with an exhausted and immunosuppressive tumor microenvironment. In summary, this study has unveiled a potential regulatory network among immune cells and the clonal dynamics of their functions, and the mechanisms of exhaustion and memory conversion between GZMK + Tem and TNFRSF13B + Bmem from antigen presentation and co-stimulation perspectives during neoadjuvant PD-1 blockade treatment in ESCC.

PDZK1 improves ventricular remodeling in hypertensive rats by regulating the stability of the Mas receptor.

Ventricular remodeling is one of the main causes of mortality from heart failure due to hypertension. Exploring its mechanism and finding therapeutic targets have become urgent scientific problems to be solved. A number of studies have shown that Mas, as an Ang-(1-7) specific receptor, was significantly reduced in myocardial tissue of rats undergoing hypertensive ventricular remodeling. It has been reported that Mas receptor levels are significantly downregulated in myocardium undergoing ventricular remodeling, but studies focused on intracellular and post-translational modifications of Mas are lacking. The results of this research are as follows: (1) PDZK1 interacts with the carboxyl terminus of Mas through its PDZ1 domain; (2) the expression of PDZK1 and Mas is decreased in rats undergoing hypertensive ventricular remodeling, and PDZK1 upregulation can ameliorate hypertensive myocardial fibrosis and myocardial hypertrophy; (3) PDZK1 enhances the stability of Mas protein through the proteasome pathway, and the proteasome inhibitor MG132 promotes hypertensive ventricular remodeling. PDZK1 improves ventricular remodeling in hypertensive rats by regulating Mas receptor stability. This study provides a scientific basis for the prevention and treatment of ventricular remodeling.

The Typical Polybrominated Diphenyl Ethers (PBDEs) and Heavy Metals Distributions in a Formal e-Waste Dismantling Site.

Understanding the release of pollutants from the formal e-waste dismantling site could provide the basic information and potential risk to guide the normative regulation of the process. In this study, the distribution of typical polybrominated diphenyl ethers (PBDEs) and heavy metals in a relocating site of a formal e-waste dismantling company was firstly investigated down to the saturated zone, with a maximum depth of 3.0 m. The mean concentrations of Σ13PBDEs were ranged from 2.815 to 7.178 ng/g, with a peak value of 7.178 ng/g in storage area. BDE-209 was the predominant congener of PBDEs in the soil, with the value ranged from 1.688 to 2.483 ng/g. A higher pollution of PBDEs and HMs was presented in the storage area. The risk assessment of PBDEs mostly posed a low environmental risk (RQ ≤ 0.01) and pentaBDE was found to be the most harmful driver for the potential environmental risk.

Association of Pathologic Complete Response and Long-Term Survival Outcomes Among Patients Treated With Neoadjuvant Chemotherapy or Chemoradiotherapy for NSCLC: A Meta-Analysis.

Introduction: Increased efforts to optimize outcomes for early stage NSCLC through the investigation of novel perioperative treatment strategies are ongoing. An emerging question is the role of pathologic response and its association with long-term clinical outcomes after neoadjuvant therapy. Methods: To investigate the association of pathologic complete response (pCR) and event-free survival (EFS) and overall survival (OS), we performed a systematic review and meta-analysis identifying studies reporting on the prognostic impact of pCR after neoadjuvant chemotherapy or chemoradiotherapy. To evaluate this prognostic value, an aggregated data (AD) meta-analyses was conducted to estimate the pooled hazard ratios (HRs) of EFS and OS for pCR. Using reconstructed individual patient data (IPD), pooled Kaplan-Meier curves were obtained to estimate this association in a more granular fashion. Subgroup analyses were conducted to further explore the impacts of study-level characteristics. Results: A total of 28 studies comprising 7011 patients were included in the AD meta-analysis, of which, IPD was available for 6274 patients from 24 studies. Results from our AD meta-analysis revealed a pooled pCR rate of 18% (95% confidence interval [CI]: 15%-21%), including significant improvements in OS (HR = 0.50, 95% CI: 0.45-0.56) and EFS (HR = 0.46, 95% CI: 0.37-0.57) on the basis of pCR status. Our IPD analysis revealed a 5-year OS rate of 63% (95% CI: 59.6-67.4) for patients with a pCR compared with 39% (95% CI: 34.5-44.5) for those without a pCR. Conclusions: pCR after neoadjuvant chemotherapy plus or minus radiotherapy is associated with significant improvements in EFS and survival for patients with resectable NSCLC.

Integrative tumour mutation burden with CD39 and PD-L1 for the prediction of response to PD-L1 blockade and adjuvant chemotherapy in muscle-invasive bladder cancer patients.

BACKGROUND: CD39, a rate-limiting enzyme to convert extracellular ATP (eATP) to adenosine, has been reported to be a key modulator of immune response, but its correlation with therapeutic sensitivity remains obscure. We conducted this study to determine whether the integration of CD39 and traditional biomarkers could improve the prediction of responsiveness to PD-L1 blockade and platinum-based chemotherapy. METHODS: We retrospectively enrolled a total of 760 patients from IMvigor210 trial, TCGA database and Zhongshan Hospital in this study. We constructed the CPT scoring system based on CD39, PD-L1 and tumour mutation burden (TMB) and validated its efficacy in predicting therapeutic responsiveness in MIBC patients. Kaplan-Meier survival and Cox regression analyses were applied to assess clinical outcomes of patients. RESULTS: The CPT scoring system could predict the response to PD-L1 blockade and platinum-based chemotherapy. The CPT score was positively correlated with APOBEC mutational signature and SNV neoantigens enrichment, antigen presentation, and TCR signalling. High CPT score also indicated the inflamed immune phenotype and basal/squamous molecular subtype. CONCLUSIONS: CD39 expression is closely correlated with the immunogenic contexture of MIBC. Integrating CD39 with PD-L1 and TMB could stratify the sensitivity of patients with MIBC to PD-L1 blockade and platinum-based chemotherapy.

CD103+CD8+ tissue-resident memory T cell infiltration predicts clinical outcome and adjuvant therapeutic benefit in muscle-invasive bladder cancer.

BACKGROUND: CD103+CD8+ tissue-resident memory T (TRM) cells, associated with better overall survival among various malignancies, are thought to activate anti-tumour immune response and affect therapeutic sensitivity including both immunotherapy and adjuvant chemotherapy (ACT). METHODS: Totally 650 muscle-invasive bladder cancer (MIBC) patients from three independent cohorts were included in this study for survival and cisplatin-based ACT response analysis. Another public data set consisting of 195 patients from IMvigor210 trial receiving PD-L1 blockade were involved in the assessment of immunotherapeutic response. Fifty-nine fresh tumour tissues were used to evaluate immune infiltration of CD103+CD8+ TRM cells. RESULTS: Patients with high CD103+CD8+ TRM cells infiltration, but not CD8+ T cells, are more likely to benefit from immunotherapy and ACT. The presence of TRM cells is highly associated with an enhanced IFNγ-enriched and T cell-inflamed anti-tumour microenvironment. Elevated CD103+CD8+ TRM cells infiltration correlated with superior ACT response in mismatch repair (MMR), homologous recombination (HR), PIK3CA/AKT and RAS/RAF pathway proficient or histone modification and cell cycle pathway deficient patients. CONCLUSIONS: CD103+CD8+ TRM cells played a crucial role in anti-tumour immunity and served as an ideal prognostic biomarker. It could be treated as a superior companion predictor for treatment response to PD-L1 inhibitor and ACT within MIBC patients.

Stromal Tumor-Associated Macrophage Infiltration Predicts Poor Clinical Outcomes in Muscle-Invasive Bladder Cancer Patients.

BACKGROUND: This study aims to reveal the clinical significance of stromal-infiltrating tumor-associated macrophages (TAMs) in muscle-invasive bladder cancer (MIBC). PATIENTS AND METHODS: This study included 288 patients from the TCGA database and 118 patients from Fudan University Shanghai Cancer Center with MIBC. The CIBERSORT model and immunohistochemistry were used to evaluate TAM infiltration. Cox regression analyses were employed to calculate their prognostic value. RESULTS: Among all 23 immune phenotypes analyzed in the TCGA cohort, pan-macrophage infiltration was significantly associated with poor prognosis (p = 0.001). Further analyses found that stromal TAM infiltration could be an independent prognostic predictor for recurrence-free survival (RFS; HR: 1.019, 95% CI: 1.006-1.033, p = 0.004). High stromal infiltration was related to unfavorable RFS. After stratification by adjuvant chemotherapy (ACT), patients without ACT could be differentiated by TAM infiltration (p = 0.036), while patients with ACT could not. Moreover, TAM infiltration was negatively associated with IFN-γ-related mRNA panel, which was shown to have strong predictive value for clinical response to programmed death-1 (PD-1) inhibition. CONCLUSIONS: Stromal TAM infiltration could be an independent prognosticator for MIBC patients. This might have potential to guide precise treatments such as ACT and immune checkpoint blockade in MIBC.

TIGIT and PD-1 expression atlas predicts response to adjuvant chemotherapy and PD-L1 blockade in muscle-invasive bladder cancer.

BACKGROUND: TIGIT and PD-1 are checkpoint receptors that could regulate the functional status of immune cells through independent pathways. However, the clinical significance of immune classification based on TIGIT and PD-1 expression remains unclear in muscle-invasive bladder cancer (MIBC). METHODS: Patients with MIBC from four independent cohorts were categorised into three clusters. Survival analysis conducted through Kaplan-Meier curves and Cox regression model. Immune contexture was measured by immunohistochemistry and CIBERSORT algorithm. Twenty-five fresh tumour tissue samples were utilised to evaluate functional state of CD8+ T cells by flow cytometry. RESULTS: Cluster I (TIGITlowPD-1low) contained widely poor immune infiltrates with higher FGFR3 mutation, Cluster II (TIGITlowPD-1high) exhibited a highly infiltrated contexture with increased cytolytic CD8+ T cells and had the best prognosis, Cluster III (TIGIThigh) presented a suppressive tumour microenvironment (TME) featured by exhausted CD8+ T cells and basal molecular subtype. Patients of Cluster III had the worst survival but could benefit more from adjuvant chemotherapy and anti-PD-L1 immunotherapy, and also presented limited FGFR3 signalling signature but activated immunotherapeutic and EGFR-associated pathway. CONCLUSIONS: TIGIT/PD-1-based risk stratification with distinct immune and molecular features could be served as a predictor for systematic therapeutic response including adjuvant chemotherapy and immunotherapy in MIBC patients.

Immune inactivation by neuropilin-1 predicts clinical outcome and therapeutic benefit in muscle-invasive bladder cancer.

BACKGROUND: Immune checkpoint blockade (ICB) and adjuvant chemotherapy (ACT) have shown clinical benefit in muscle-invasive bladder cancer (MIBC) with only a few predictive biomarkers identified so far. Neuropilin-1 (NRP1) has been identified as a key immune checkpoint and a novel immunotherapeutic target but the clinical significance of NRP1 remains unclear in MIBC. METHODS: Three independent cohorts were involved in our study: IMvigor210 Cohort (n = 348), The Cancer Genome Atlas Cohort (TCGA, n = 391), and Zhongshan Hospital Cohort (ZSHS, n = 130). Parallel detection and validation of risk stratification based on NRP1 expression were executed in patients treated with anti-PD-L1 agent and adjuvant chemotherapy (ACT). RESULTS: NRP1 expression conferred poor survival and predicted response to both PD-L1 blockade and cisplatin-based ACT in MIBC. Further exploration revealed high-level NRP1 was extremely associated with infiltration of exhausted CD8+ T cells, immature NK cells and M2 polarized tumor-associated macrophages in MIBC patients. Moreover, elevated NRP1 expression was also correlated with low mutation burden and reduced mutation in cell cycle pathway. CONCLUSIONS: Our study firstly identified and validated the clinical implications of NRP1 expression for prognosis and systematic therapeutic responses (PD-L1 blockade and ACT) in MIBC. NRP1 expression was associated with an immunosuppressive microenvironment with dysfunctional effector immune cells. Prospective investigations of its roles in the therapeutic landscape of MIBC warrant more consideration.

Infiltration and Polarization of Tumor-associated Macrophages Predict Prognosis and Therapeutic Benefit in Muscle-Invasive Bladder Cancer.

BACKGROUND: Muscle-invasive bladder cancer (MIBC) is an aggressive and heterogeneous malignancy. Tumor-associated macrophages (TAMs) are key infiltrating cell populations in the inflammatory microenvironment of malignant tumors including MIBC. It intrigues us to explore the clinical significance and immunoregulatory role of TAMs infiltration and polarization in MIBC. METHODS: A total of 141 patients with MIBC from Zhongshan Hospital and 391 patients with MIBC from The Cancer Genome Atlas (TCGA) database were included in this study. Moreover, 195 patients who received anti-PD-L1 therapy from the IMvigor210 trial were enrolled. Patients were categorized into three subtypes considering the infiltration level and polarization status of TAMs, denoted as TAMlow (Subtype I), TAMhigh&M2/M1low (Subtype II), and TAMhigh&M2/M1high (Subtype III). RESULTS: Subtype III suffered inferior prognosis, and Subtype II could benefit more from adjuvant chemotherapy (ACT). Subtype III was featured with increased pro-tumor cells and immunosuppressive cytokines, while Subtype II possessed more immunogenic cells infiltration with activated and tumoricidal properties. Subtype II and Subtype III presented basal/squamous-like characterization and showed additional prognostic merit beyond molecular classification. Subtype I exhibited elevated level of FGFR3 signature, while Subtype II had EGFR signaling activation and immunotherapeutic indication. Additionally, Subtype II patients were indeed highly sensitive to PD-L1 blockade therapy in IMvigor210 trial. CONCLUSION: The infiltration and polarization status of TAMs shaped distinct immune microenvironment with predictive significance for survival outcome, ACT benefit, and PD-L1 blockade therapy sensitivity in MIBC. Immune classification based on TAMs polarization and infiltration might provide tools to tailor chemotherapy and immunotherapy.

Latency-associated peptide identifies therapeutically resistant muscle-invasive bladder cancer with poor prognosis.

BACKGROUND: Latency-associated peptide (LAP) was identified as crucial immune regulator in tumor microenvironment (TME) in recent researches. In this study, we aimed to estimate the predictive value of LAP expression for clinical survival and therapeutic response in muscle-invasive bladder cancer (MIBC). METHODS: Our study encompassed 140 MIBC patients from Zhongshan Hospital (ZSHS cohort), 401 patients from The Cancer Genome Atlas (TCGA cohort) and 195 patients received PDL1 blockade from IMvigor210 trial. Survival analyses were conducted through Kaplan-Meier curve and Cox regression model. LAP expression and its association with immune contexture were evaluated in ZSHS and TCGA cohort. RESULTS: We found that high intratumoral LAP+ cells infiltration anticipated inferior survival and adjuvant chemotherapy (ACT) response, and was closely related to an immunoevasive contexture with increased M2 macrophages, neutrophils and conspicuously a cluster of highly exhausted CD8+ T cells. The combinational analysis of LAP+ cells and CD8+ T cells infiltration stratified patients into distinct risk groups with implications for therapeutic sensitivity to PDL1 blockade and refinement of molecular classification in MIBC. CONCLUSIONS: LAP expression was correlated with patients' inferior prognosis, ACT-tolerance and an immunoevasive TME with exhausted CD8+ T cell infiltration, suggesting that LAP could serve as a promising therapeutic target in MIBC. Simultaneously, our novel TME classification based on LAP+ cells and CD8+ T cells infiltration and its potential in appraising PDL1 blockade application for MIBC patients deserved further validation.

Nox4-dependent ROS production is involved in CVB3-induced myocardial apoptosis.

Viral myocarditis is a cardiovascular disease that seriously affects human health. Its mechanism is not clear. Coxsackievirus B3 (CVB3) is a member of the picornavirus family and is the leading cause of viral myocarditis. Our group tested the genes in a mouse model of CVB3 virus infection and confirmed that the NADPH oxidase gene had a high expression trend in the acute phase of infection. Whether Nox4, the homologue of NADPH oxidase, participates in the process of viral myocarditis has not been reported. In this study, we found increased expression of Nox4 in viral myocarditis in vivo and in vitro. DPI is a non-specific inhibitor of Nox4 that improved CVB3-induced myocarditis after injection in vivo. DPI also inhibited intracellular ROS release and apoptosis in vitro. Our data indicated that Nox4-dependent ROS production was involved in CVB3-induced myocardial apoptosis.

Involvement of mitochondrial fission in calcium sensing receptor-mediated vascular smooth muscle cells proliferation during hypertension.

Hyperproliferation of vascular smooth muscle cells (VSMC) is a major risk factor for cardiovascular diseases. Proper mitochondrial fission and fusion is involved with VSMC function. However, the role and mechanism of mitochondrial morphological changes in VSMC proliferation are not well understood. Here, we found that calcium sensing receptor (CaSR) was increased in the aortas from spontaneous hypertensive rats (SHRs) compared with age-matched Wistar Kyoto (WKY) rats. There was also an increase in mitochondrial fission and VSMC proliferation, which was attenuated by Calhex231. In primary rat VMSC, angiotensin II (Ang II) stimulation induced cytosolic [Ca2+]i increase, mitochondrial shortening and proliferation, all of which could be attenuated by pretreatment with mitochondrial division inhibitor-1 (Mdivi-1) and Calhex231. Our data indicate that CaSR-mediated mitochondrial fission could be a therapeutic target for hyperproliferative disorders.

PEITC in End-Stage B-Cell Prolymphocytic Leukemia: Case Report of Possible Sensitization to Salvage R-CHOP.

INTRODUCTION: B-cell prolymphocytic leukemia (B-PLL) is a rare, aggressive leukemia distinct from chronic lymphocytic leukemia, with median survival of only 3 years. B-PLL is resistant to most chemotherapy and newer targeted therapies such as alemtuzumab and thalidomide. Phenylethyl isothiocyanate (PEITC) is a natural compound from horseradish with evidence for therapeutic potential in multiple leukemia types. CASE PRESENTATION: Here we present a case report of a 53-year-old man whose chronic lymphocytic leukemia transformed to end-stage B-PLL, disqualifying him for allogenic stem cell transplantation. He was treated with PEITC followed by salvage R-CHOP (Rituximab, Cyclophosphamide, Hydroxydaunorubicin [doxorubicin hydrochloride], Oncovin [vincristine sulfate], Prednisone or Prednisolone) chemotherapy, which led to normalized white blood cell count and disease stabilization that requalified him for allogenic peripheral stem-cell transplant therapy. We conducted a systematic review to analyze and interpret the potential contribution of PEITC to his unexpectedly favorable R-CHOP response. Following sequential 8 weeks of PEITC/pentostatin and 6 cycles of R-CHOP, the patient received allogenic peripheral blood stem cell transplant on an outpatient basis and remains well at the time of this publication, with no evidence of CD20+ small B-cells. DISCUSSION: Given the limited data for R-CHOP in B-PLL, this patient's recovery suggests presensitization of B-PLL cells toward R-CHOP, potentially justifying further investigation.

Warm-white light-emitting diode with high color rendering index fabricated by combining trichromatic InGaN emitter with single red phosphor.

We present a trichromatic GaN-based light-emitting diode (LED) that emits near-ultraviolet (n-UV) blue and green peaks combined with red phosphor to generate white light with a low correlated color temperature (CCT) and high color rendering index (CRI). The LED structure, blue and green unipolar InGaN/GaN multiple quantum wells (MQWs) stacked with a top p-i-n structure containing an InGaN/GaN MQW emitting n-UV light, was grown epitaxially on a single substrate. The trichromatic LED chips feature a vertical conduction structure on a silicon substrate fabricated through wafer bonding and laser lift-off techniques. The blue and green InGaN/GaN MQWs were pumped with n-UV light to re-emit low-energy photons when the LEDs were electrically driven with a forward current. The emission spectrum included three peaks at approximately 405, 468, and 537 nm. Furthermore, the trichromatic LED chips were combined with red phosphor to generate white light with a CCT and CRI of approximately 2900 and 92, respectively.

GaN-based photon-recycling green light-emitting diodes with vertical-conduction structure.

A p-i-n structure with near-UV(n-UV) emitting InGaN/GaN multiple quantum well(MQW) structure stacked on a green unipolar InGaN/GaN MQW was epitaxially grown at the same sapphire substrate. Photon recycling green light-emitting diodes(LEDs) with vertical-conduction feature on silicon substrates were then fabricated by wafer bonding and laser lift-off techniques. The green InGaN/GaN QWs were pumped with n-UV light to reemit low-energy photons when the LEDs were electrically driven with a forward current. Efficiency droop is potentially insignificant compared with the direct green LEDs due to the increase of effective volume of active layer in the optically pumped green LEDs, i.e., light emitting no longer limited in the QWs nearest to the p-type region to cause severe Auger recombination and carrier overflow losses.

Novel active personal nanoparticle sampler for the exposure assessment of nanoparticles in workplaces.

A novel active personal nanoparticle sampler (PENS), which enables the collection of both respirable particulate mass (RPM) and nanoparticles (NPs) simultaneously, was developed to meet the critical demand for personal sampling of engineered nanomaterials (ENMs) in workplaces. The PENS consists of a respirable cyclone and a micro-orifice impactor with the cutoff aerodynamic diameter (d(pa50)) of 4 µm and 100 nm, respectively. The micro-orifice impactor has a fixed micro-orifice plate (137 nozzles of 55 µm in the inner diameter) and a rotating, silicone oil-coated Teflon filter substrate at 1 rpm to achieve a uniform particle deposition and avoid solid particle bounce. A final filter is used after the impactor to collect the NPs. Calibration results show that the d(pa50) of the respirable cyclone and the micro-orifice impactor are 3.92 ± 0.22 µm and 101.4 ± 0.1 nm, respectively. The d(pa50) at the loaded micro-Al(2)O(3) mass of 0.36-3.18 mg is shifted to 102.9-101.2 nm, respectively, while it is shifted to 98.9-97.8 nm at the loaded nano-TiO(2) mass of 0.92-1.78 mg, respectively. That is, the shift of d(pa50) due to solid particle loading is small if the PENS is not overloaded. Both NPs and RPM concentrations were found to agree well with those of the IOSH respirable cyclone and MOUDI. By using the present PENS, the collected samples can be further analyzed for chemical species concentrations besides gravimetric analysis to determine the actual exposure concentrations of ENMs in both RPM and NPs fractions in workplaces, which are often influenced by the background or incident pollution sources.