Novel artesunate-metformin conjugate inhibits bladder cancer cell growth associated with Clusterin/SREBP1/FASN signaling pathway.

Bladder cancer remains the 10th most common cancer worldwide. In recent years, metformin has been found to have potential anti-bladder cancer activity while high concentration of IC50 at millimolar level is needed, which could not be reached by regular oral administration route. Thus, higher efficient agent is urgently demanded for clinically treating bladder cancer. Here, by conjugating artesunate to metformin, a novel artesunate-metformin dimer triazine derivative AM2 was designed and synthesized. The inhibitory effect of AM2 on bladder cancer cell line T24 and the mechanism underlying was determined. Anti-tumor activity of AM2 was assessed by MTT, cloning formation and wound healing assays. Decreasing effect of AM2 on lipogenesis was determined by oil red O staining. The protein expressions of Clusterin, SREBP1 and FASN in T24 cells were evaluated by Western blotting. The results show that AM2 significantly inhibited cell proliferation and migration at micromolar level, much higher than parental metformin. AM2 reduced lipogenesis and down-regulated the expressions of Clusterin, SREBP1 and FASN. These results suggest that AM2 inhibits the growth of bladder cancer cells T24 by inhibiting cellular lipogenesis associated with the Clusterin/SREBP1/FASN signaling pathway.

ARF4-mediated retrograde trafficking as a driver of chemoresistance in GBM.

BACKGROUND: Cellular functions hinge on the meticulous orchestration of protein transport, both spatially and temporally. Central to this process is retrograde trafficking, responsible for targeting proteins to the nucleus. Despite its link to many diseases, the implications of retrograde trafficking in glioblastoma (GBM) are still unclear. METHODS: To identify genetic drivers of TMZ resistance, we conducted comprehensive CRISPR-knockout screening, revealing ADP-ribosylation factor 4 (ARF4), a regulator of retrograde trafficking, as a major contributor. RESULTS: Suppressing ARF4 significantly enhanced TMZ sensitivity in GBM patient-derived xenograft (PDX) models, leading to improved survival rates (p<0.01) in both primary and recurrent lines. We also observed that TMZ exposure stimulates ARF4-mediated retrograde trafficking. Proteomics analysis of GBM cells with varying levels of ARF4 unveiled the influence of this pathway on EGFR signaling, with increased nuclear trafficking of EGFR observed in cells with ARF4 overexpression and TMZ treatment. Additionally, spatially-resolved RNA-sequencing of GBM patient tissues revealed substantial correlations between ARF4 and crucial nuclear EGFR (nEGFR) downstream targets, such as MYC, STAT1, and DNA-PK. Decreased activity of DNA-PK, a DNA repair protein downstream of nEGFR signaling that contributes to TMZ resistance, was observed in cells with suppressed ARF4 levels. Notably, treatment with DNA-PK inhibitor, KU57788, in mice with a recurrent PDX line resulted in prolonged survival (p<0.01), highlighting the promising therapeutic implications of targeting proteins reliant on ARF4-mediated retrograde trafficking. CONCLUSION: Our findings demonstrate that ARF4-mediated retrograde trafficking contributes to the development of TMZ resistance, cementing this pathway as a viable strategy to overcome chemoresistance in GBM.

Risk factors for the development of oral precancerous lesions in a cohort of 293 oral lichen planus patients with or without chronic periodontitis in southern Taiwan.

Background/purpose: Oral lichen planus (OLP) may contribute to the risk of chronic periodontitis, and no reports have shown whether OLP patients with periodontitis have a greater risk of oral precancerous lesions, Candida infection or other clinicopathological diseases. This study aimed to assess the risk factors for the development of oral precancerous lesions in a cohort of 293 OLP patients with or without chronic periodontitis in southern Taiwan. Materials and methods: The current study recruited 293 OLP patients without preexisting periodontitis at a tertiary institution from 1995 to 2018. The patients were divided into two groups based on the presence or absence of periodontitis. The study compared various clinical and pathological characteristics between the two groups, and also estimated the odds ratio (OR) and the 10-year cumulative risk of chronic periodontitis in OLP patients using logistic regression models and Kaplan‒Meier analysis methods, respectively. Results: After adjusting for age and gender, the significant contributors to oral precancerous lesions in OLP patients (P < 0.05) were periodontal disease (OR = 2.24) and the male gender (OR = 7.52). Betel nut consumption (OR = 2.61), smoking (OR = 2.46), and candidiasis infection (OR = 3.02) also showed significant associations. Older OLP patients had a lower lesion risk, while a longer OLP duration heightened the periodontal disease likelihood. Conclusion: The present study demonstrated that coexisting periodontal disease increases the likelihood of developing precancerous lesions in patients with OLP. Periodontal management with oral hygiene care and quitting betel nut consumption and smoking can reduce the risk.

Pseudopterygia in Fuchs Superficial Marginal Keratitis: Clinical Course and Surgical Outcome.

PURPOSE: Surgery for pseudopterygia in Fuchs superficial marginal keratitis (FSMK) bears the risk of corneal perforation, as described in a few case reports. The aim of this case series was to understand the clinical course and surgical outcomes of pseudopterygia in FSMK. METHODS: A retrospective case series included patients meeting FSMK criteria with pseudopterygia in at least 1 eye. The severity grading of pseudopterygia and peripheral infiltration events at follow-up were analyzed. Pseudopterygia involving corneal central 3 mm diameter (grade III) received surgery. Peripheral corneal infiltrate events within 1 week after surgery were recorded. RESULTS: Thirty-three eyes of 19 patients (8 men, 11 women; age 40-85 years; mean, 65 years) were included, with an average 48.1-month follow-up (range 0-188.8 months). At presentation, 7 eyes (21%) had grade III pseudopterygia. One patient showed corneal perforation 3 days after "pterygium" surgery elsewhere. The contralateral eye met the diagnosis of FSMK. During follow-up, 16 eyes (49%) exhibited peripheral corneal infiltrates, and 7 eyes (21%) showed progression of pseudopterygia to higher grades before or without any surgery. Ten eyes with grade III pseudopterygia underwent surgery. The best-corrected visual acuity in Logarithm of the Minimum Angle of Resolution improved from 0.34 ± 0.18 (range 0.05-0.52) to 0.13 ± 0.11 (range 0.05-0.4) (P = 0.0023). Six eyes (60%) developed peripheral corneal infiltrates within 1 week after surgery, which responded well to topical corticosteroids. CONCLUSIONS: Pseudopterygia in FSMK may progress to threaten visual acuity. Surgical excision can be safe and can effectively improve vision on the condition that the patients are closely followed in the early postoperative period to notice the corneal infiltrates.

Novel Manifestation of Corneal Dystrophy After Keratorefractive Surgery.

PURPOSE: This study aimed to report cases of bilateral corneal Bowman layer deposits in 4 patients with a history of keratorefractive surgery. To our knowledge, this condition has not previously been reported and should be distinguished from granular corneal dystrophy type 2 and other corneal dystrophies. METHODS: We reviewed all available medical records that were collected between January 2010 and December 2021 at a tertiary referral center and performed whole-exome sequencing to provide diagnostic information. RESULTS: Four patients exhibited similar bilateral corneal deposits that were observed more than 10 years after keratorefractive surgery. The patients' ages ranged from 36 to 53 years; 3 of the 4 patients were female. Three patients received laser in situ keratomileusis surgery, and 1 received radial keratotomy. All 4 patients denied having a family history of ocular diseases and reported an uneventful postoperative course. On examination, the best-corrected visual acuity ranged from 6/10 to 6/6 in all 4 patients. Slit-lamp examination revealed bilateral superficial corneal deposits involving the central cornea, and anterior segment optical coherence tomography revealed hyperreflective deposits located in the Bowman layer. Such unique manifestations suggested corneal dystrophy; thus, whole-exome sequencing was performed on all 4 patients. Only 1 patient exhibited a missense mutation in TGFBI . We further analyzed common de novo mutations to explore possible candidate genes associated with this presentation. CONCLUSIONS: We report a rare entity of presumed corneal dystrophy with deposits located in the Bowman layer in 4 patients who had received keratorefractive surgery. Clarifying the underlying pathophysiology and genetic predisposition of this disease may aid in diagnosing and preventing potential complications after keratorefractive surgery.

Estimating genome-wide DNA methylation heterogeneity with methylation patterns.

BACKGROUND: In a heterogeneous population of cells, individual cells can behave differently and respond variably to the environment. This cellular diversity can be assessed by measuring DNA methylation patterns. The loci with variable methylation patterns are informative of cellular heterogeneity and may serve as biomarkers of diseases and developmental progression. Cell-to-cell methylation heterogeneity can be evaluated through single-cell methylomes or computational techniques for pooled cells. However, the feasibility and performance of these approaches to precisely estimate methylation heterogeneity require further assessment. RESULTS: Here, we proposed model-based methods adopted from a mathematical framework originally from biodiversity, to estimate genome-wide DNA methylation heterogeneity. We evaluated the performance of our models and the existing methods with feature comparison, and tested on both synthetic datasets and real data. Overall, our methods have demonstrated advantages over others because of their better correlation with the actual heterogeneity. We also demonstrated that methylation heterogeneity offers an additional layer of biological information distinct from the conventional methylation level. In the case studies, we showed that distinct profiles of methylation heterogeneity in CG and non-CG methylation can predict the regulatory roles between genomic elements in Arabidopsis. This opens up a new direction for plant epigenomics. Finally, we demonstrated that our score might be able to identify loci in human cancer samples as putative biomarkers for early cancer detection. CONCLUSIONS: We adopted the mathematical framework from biodiversity into three model-based methods for analyzing genome-wide DNA methylation heterogeneity to monitor cellular heterogeneity. Our methods, namely MeH, have been implemented, evaluated with existing methods, and are open to the research community.

Ribonucleotide reductase regulatory subunit M2 drives glioblastoma TMZ resistance through modulation of dNTP production.

During therapy, adaptations driven by cellular plasticity are partly responsible for driving the inevitable recurrence of glioblastoma (GBM). To investigate plasticity-induced adaptation during standard-of-care chemotherapy temozolomide (TMZ), we performed in vivo single-cell RNA sequencing in patient-derived xenograft (PDX) tumors of GBM before, during, and after therapy. Comparing single-cell transcriptomic patterns identified distinct cellular populations present during TMZ therapy. Of interest was the increased expression of ribonucleotide reductase regulatory subunit M2 (RRM2), which we found to regulate dGTP and dCTP production vital for DNA damage response during TMZ therapy. Furthermore, multidimensional modeling of spatially resolved transcriptomic and metabolomic analysis in patients' tissues revealed strong correlations between RRM2 and dGTP. This supports our data that RRM2 regulates the demand for specific dNTPs during therapy. In addition, treatment with the RRM2 inhibitor 3-AP (Triapine) enhances the efficacy of TMZ therapy in PDX models. We present a previously unidentified understanding of chemoresistance through critical RRM2-mediated nucleotide production.

Synthesis and Antitumor Activities of Novel Mitochondria-Targeted Dihydroartemisinin Ether Derivatives.

Ten novel mitochondria-targeted dihydroartemisinin ether derivatives were designed, synthesized, and evaluated for antitumor activity against five cancer cell lines in vitro. Profoundly, compound D8-T (IC50 = 56.9 nM) showed the most potent antiproliferative activity against the T24 cells with low cytotoxicity in normal human umbilical vein endothelial cells. High-performance liquid chromatography analysis confirmed that D8-T targeted mitochondria 6.3-fold higher than DHA. ATP content assay demonstrated that D8-T decreased the ATP level of bladder cancer cells. The effect of D8-T on cell apoptosis was determined by flow cytometry and western blot of Bax and Bcl-2. Surprisingly, the results indicated that D8-T did not induce bladder cancer cell apoptosis. In contrast, the cell cycle analysis and western blot of CDK4, CDK6, cyclin D1, and p21 demonstrated that the cancer cell cycle was arrested at the G1 phase after D8-T treatment. Furthermore, the consistent results were received by RNA-seq assay. These promising findings implied that D8-T could serve as a great candidate against bladder cancer for further investigation.

Cataract surgery in patients with Fuchs' dystrophy and corneal decompensation indicated for Descemet's membrane endothelial keratoplasty.

The availability of corneal donor tissue is limited in most developing countries. This study evaluated whether patients with coexisting cataract and Fuchs' dystrophy with corneal decompensation awaiting Descemet's membrane endothelial keratoplasty (DMEK) benefited from phacoemulsification. This is a retrospective case-control study which included patients with Fuchs' dystrophy and evidence of corneal decompensation awaiting DMEK. Best-corrected visual acuity (BCVA) and central corneal thickness (CCT) were documented at baseline (pre-cataract surgery in the case group, or at the time of transplantation registry in the control group), 1-month and pre-DMEK. A total of 16 phakic patients with visually significant cataracts had cataract surgery during the study period, and 15 pseudophakic patients were included as controls. There was no significant difference with regard to BCVA at baseline, 1-month or pre-DMEK between the case and control groups. Similarly, no significant difference in CCT was found at baseline, 1-month or pre-DMEK. In the case group, 4 patients with improved visual acuity post-cataract surgery chose to defer DMEK. After stratification, statistical analysis showed significantly better BCVA in the deferred group (n = 4) at 1-month post-cataract surgery, compared to the DMEK group (n = 12) (0.21 ± 0.21 vs. 0.86 ± 0.29 LogMAR, P = 0.004). The other parameters, including baseline BCVA and CCT at any time point documented, were not statistically different. In conclusion, in patients with Fuchs' dystrophy and decompensated corneas awaiting transplantation, phacoemulsification did not lead to significant increase of corneal thickness nor deterioration of visual acuity. A few patients achieved satisfactory vision after cataract surgery and deferred endothelial keratoplasty.

Novel dihydroartemisinin derivative Mito-DHA5 induces apoptosis associated with mitochondrial pathway in bladder cancer cells.

BACKGROUND: Bladder cancer is the second most common genitourinary malignancy and the eleventh most common cancer worldwide. Dihydroartemisinin (DHA), a first-line antimalarial drug, has been found to have potent antitumor activity. In our previous study, a novel dihydroartemisinin derivative Mito-DHA5 synthesized in our laboratory has a stronger anti-tumor activity than DHA. In this study, we investigated the apoptotic effect of Mito-DHA5 on bladder cancer T24 cells and molecular mechanisms underlying. METHODS: Antitumor activity in vitro was evaluated by MTT, wound healing and cloning formation assays. Mitochondrial membrane potential (MMP) was detected by JC-1 probe and ROS levels were measured by specific kit. The expression of caspase-3, cleaved-caspase3, mitochondrial Cyt-C, Bcl-2, Bax and PARP in T24 cells was evaluated by Western blotting. RESULTS: The results showed that Mito-DHA5 reduced cell viability with an IC50 value of 3.2 µM and induced T24 cell apoptosis in a dose-dependent manner, increased the production of ROS and decreased MMP. Mito-DHA5 could down-regulate the expression of Bcl-2, mitochondrial Cyt-C, Caspase-3, PARP and up-regulate the expression of Bax and cleaved Caspase-3. CONCLUSIONS: These data suggested that Mito-DHA5 had a potent inhibitory effect on T24 bladder cancer cell growth and induced these cells apoptosis associated with mitochondrial pathway.

Comparison of clinical outcomes of LASIK, Trans-PRK, and SMILE for correction of myopia.

Transepithelial photorefractive keratectomy (Trans-PRK), laser-assisted in situ keratomileusis (LASIK), and small incision lenticule extraction (SMILE) are three mainstay refractive surgeries worldwide. The applicability, efficacy, safety, and predictability of these different techniques are quite similar. Trans-PRK has the strongest biostability, earliest return to normal corneal sensitivity but the longest recovery time, most uncomfortable postoperative experience, and possibility of corneal haze. LASIK possesses the fastest visual rehabilitation but the slowest corneal nerve reinnervation, and flap displacement is possibly lifelong. SMILE incurs no flap-related complications and has intermediate vision recovery time and biomechanics compared with Trans-PRK and LASIK. However, it lacks the cyclotorsion-compensation system, eye-tracking system, and customized treatment profile for high astigmatism or irregular corneal surface. This review aims to introduce the mechanisms, pros, and cons of these three types of refractive surgery. With full understanding, practitioners could advise patients on the most suitable treatment of choice.

Bilateral corneal edema in an alcoholic male.

Pseudophakic bullous keratopathy and Fuchs' endothelial dystrophy are the two most common causes of corneal edema after cataract surgery. We report a 61-year-old alcoholic male with bilateral corneal edema that improved after his alcohol abstinence. He had uneventful bilateral cataract surgery 3 years ago and blurred vision in both eyes developed for weeks. As he had no history of endothelial dystrophy, the treatment for viral endotheliitis was used initially yet in vain. We asked him to stop alcohol and adjusted his psychiatric drugs, but he lied about stopping drinking. The corneal edema progressed, and finally, he underwent penetrating keratoplasty in his left eye 1 year later. During hospitalization for surgery, alcohol withdrawal syndrome was noted because he could not drink alcohol in our hospital. After he quit drinking for months, corneal edema in the right eye disappeared. Our case highlights that alcoholism can result in corneal edema, and stopping drinking is necessary in these patients.

Involvement of prohibitin 1 and prohibitin 2 upregulation in cBSA-induced podocyte cytotoxicity.

Membranous nephropathy (MN) is the most common cause of nephrotic syndrome in adults, when not effectively treated. The aim of this study was to discover new targets for the diagnosis and treatment of MN. A reliable mouse model of MN was used by the administration of cationic bovine serum albumin (cBSA). Mice with MN exhibited proteinuria, histopathological changes, and accumulation of immune complexes in the glomerular basement membrane. Label-free proteomics analysis was performed to identify changes in protein expression, and the overexpressed proteins were evaluated. There were 273 proteins that showed significantly different expression in mice with MN, as compared to the controls. String analysis showed that functions related to cellular catabolic processes were downregulated in MN. Among the differentially expressed proteins, prohibitin 1 (PHB1) and prohibitin 2 (PHB2) were upregulated in the kidneys of mice with MN, as demonstrated by immunohistochemistry (IHC), and this upregulation was observed in both the tubular cells and glomeruli. Both shRNA-mediated knockdown of PHB1 or PHB2 inhibited tumor suppressor p53 expression and significantly promoted podocyte proliferation. In addition, both PHB1 and PHB2 were responsible for cBSA-induced cytotoxicity. Microarray analysis further revealed that the upregulation of PHB1 and PHB2 may be due to a blockage of proteasome activity. These data demonstrate that the upregulation of PHB2 is involved in cBSA-mediated podocyte cytotoxicity, which may lead to MN development.

Weaning outcome of solid cancer patients requiring mechanical ventilation in the intensive care unit.

BACKGROUND: Whether the weaning outcome of solid cancer patients receiving mechanical ventilation (MV) in the intensive care unit (ICU) is comparable to that in non-cancer patients is unknown. The aim of this study was to compare the weaning outcomes between non-cancer patients and patients with different types of cancer. METHODS: We studied patients requiring MV during ICU stay for medical reasons between 2012 and 2014. Cancer patients were grouped into those with lung cancer (LC), head and neck cancer (HNC), hepatocellular carcinoma (HCC), and other cancers (OC). The primary endpoint was successful weaning at day 90 after the initiation of MV, and the main secondary endpoints were 28-day and 90-day mortality after ICU admission. RESULTS: Five hundred and eighteen patients with solid cancers and 1362 non-cancer patients were recruited. The rate of successful weaning at day 90 was 57.9% in cancer patients, which was lower than 68.9% in non-cancer patients (p < 0.001). Compared to non-cancer patients, LC was associated with a lower probability of weaning at day 90 (hazard ratio 0.565, 95% CI 0.446 to 0.715), while HNC, HCC, and OC had similar probabilities. The 28-day and 90-day mortality rates were higher in cancer patients than in non-cancer patients (45.2% vs. 29.4%, and 65.6% vs. 37.7%, respectively, both p < 0.001). CONCLUSION: Among mechanically ventilated patients in the ICU, those with LC were associated with a lower probability of weaning at day 90 compared to non-cancer patients.

Pancreatic neuro-insular network in young mice revealed by 3D panoramic histology.

AIMS/HYPOTHESIS: It has been proposed that the neuro-insular network enables rapid, synchronised insulin secretion. However, to date, acquiring the pancreatic tissue map to study the neural network remains a challenging task as there is a lack of feasible approaches for large-scale tissue analysis at the organ level. Here, we have developed 3-dimensional (3D) panoramic histology to characterise the pancreatic neuro-insular network in young mice. METHODS: Pancreases harvested from young wild-type B6 mice (3 and 8 weeks old) and db/db mice (3 weeks old; db/db vs db/+) were used to develop 3D panoramic histology. Transparent pancreases were prepared by optical clearing to enable deep-tissue, tile-scanning microscopy for qualitative and quantitative analyses of islets and the pancreatic tissue network in space. RESULTS: 3D panoramic histology reveals the pancreatic neurovascular network and the coupling of ganglionic and islet populations via the network. This integration is identified in both 3- and 8-week-old mice, featuring the peri-arteriolar neuro-insular network and islet-ganglionic aggregation. In weaning hyperphagic db/db mice, the 3D image data identifies the associated increases in weight, adipose tissue attached to the pancreas, density of large islets (major axis > 150 µm) and pancreatic sympathetic innervation compared with db/+ mice. CONCLUSIONS/INTERPRETATION: Our work provides insight into the neuro-insular integration at the organ level and demonstrates a new approach for investigating previously unknown details of the pancreatic tissue network in health and disease.

Elevation of ß-galactoside α2,6-sialyltransferase 1 in a fructoseresponsive manner promotes pancreatic cancer metastasis.

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive type of pancreatic cancer with clinical characteristics of local invasion and early metastasis. Recent cohort studies indicate high fructose intake is associated with an increase in pancreatic cancer risk. However, the mechanisms by which fructose promotes pancreatic tumorigenesis remain unclear. Herein, Kras+/LSLG12D mice were crossed with Elas-CreER transgenic mice to determine whether fructose intake directly contributes to tumor formation. Orthotopic tumor-xenograft experiments were performed to determine whether fructose substitution enhances the metastatic potential of PDAC cells. The mechanisms underlying the effects of fructose were explored by RNAseq analysis in combination with high-performance anion exchange chromatography. Dietary fructose was initially found to promote the development of aggressive pancreatic cancer in mice conditionally expressing KrasG12D in the adult pancreas. We further revealed that fructose substitution enhanced the metastatic potential of human PDAC cell via selective outgrowth of aggressive ABCG2-positive subpopulations and elevating N-acetylmannosamine levels that upregulated ß-galactoside α2,6-sialyltransferase 1 (ST6Gal1), thereby promoting distant metastasis. Finally, we observed that PDAC patients expressing higher levels of ST6Gal1 and GLUT5 presented poorer prognosis compared to other groups. In conclusion, our findings have elucidated a crucial role of ST6Gal1 in regulating the invasiveness of PDACs in a fructose-responsive manner.

Conjunctival Acute Graft-versus-Host Disease in Adult Patients Receiving Allogeneic Hematopoietic Stem Cell Transplantation: A Cohort Study.

BACKGROUND: To investigate the incidence, risk factors and survival of conjunctival acute graft-versus-host disease (aGVHD) in adult patients undergoing allogeneic hematopoietic stem cell transplantation (HSCT). METHODS: This retrospective study included a total of 139 patients undergoing allogeneic HSCT between January 2012 and December 2014 at a tertiary referral hospital. Patients with ocular complaints after allogeneic HSCT or first donor lymphocyte infusion were evaluated by ophthalmologists. The risk factors for conjunctival aGVHD were analyzed using the Cox proportional hazards model. The overall survival was evaluated using Kaplan-Meier estimates. RESULTS: Thirteen (9.4%) patients developed conjunctival aGVHD, including eight patients with pseudomembranous conjunctivitis. The cumulative incidence of conjunctival aGVHD was 2.1 cases per 10,000 person-day. The median age at HSCT was 47 years (range, 18 to 66) in all patients and 42 years (range, 24 to 58) in the 13 patients with conjunctival aGVHD. Median time of follow-up after allogeneic HSCT was 353 days (range, 11 to 1184). In univariate analysis, grades II-IV skin aGVHD (P = 0.002) and advanced systemic aGVHD except skin aGVHD (overall grades III-IV) (P = 0.001) were significant predictors for conjunctival aGVHD. In multivariate analysis, grades II-IV skin aGVHD was a significant risk factor (P = 0.04). The severity of conjunctival aGVHD was generally correlated with the systemic aGVHD (P = 0.001). Overall survival was significantly shorter in patients with grades II-IV aGVHD compared to those with grade 0-I (P = 0.01). Survival in patients with conjunctival aGVHD did not differ significantly from those without this complication (P = 0.94). In the subgroup analysis of patients with grades III-IV aGVHD, survival was significantly longer in patients with conjunctival involvement than those without (P = 0.03). CONCLUSIONS: The severity of conjunctival aGVHD is correlated with systemic aGVHD, but not with inferior overall survival.

PanIN-associated pericyte, glial, and islet remodeling in mice revealed by 3D pancreatic duct lesion histology.

Pericytes and glial cells are accessory cells of neurovascular networks, which have been reported to participate in scar formation after tissue injury. However, it remains unclear whether similar reactive cellular responses occur in pancreatic intraepithelial neoplasia (PanIN). In this study we developed three-dimensional (3D) duct lesion histology to investigate PanIN and the associated pericyte, glial, and islet remodeling. Transparent mouse pancreata with a Kras(G12D) mutation were used to develop 3D duct lesion histology. Deep-tissue, tile-scanning microscopy was performed to generate panoramic views of the diseased pancreas for global examination of early stage and advanced duct lesion formation. Fluorescence signals of ductal and neurovascular networks were simultaneously detected to reveal associated remodeling. Significantly, in Kras(G12D)-mutant mice, when the low-grade PanINs emerge, duct lesions appear as epithelial buds with perilesional pericyte and glial activation. When PanINs occur in large scale (induced by cerulein injections to the mutant mice), the 3D image data identifies 1) aggregation of PanINs in clusters in space; 2) overexpression of the pericyte marker NG2 in the PanIN microenvironment; and 3) epithelial in-growth to islets, forming the PanIN-islet complexes. Particularly, the PanIN-islet complexes associate with proliferating epithelial and stromal cells and receive substantial neurovascular supplies, making them landmarks in the atrophic lobe. Overall, perilesional pericyte and glial activation and formation of the PanIN-islet complex underline cellular heterogeneity in the duct lesion microenvironment. The results also illustrate the advantage of using 3D histology to reveal previously unknown details of neurovascular and endocrine links to the disease.

A test protocol for assessing the hearing status of students with special needs.

OBJECTIVES: Individuals with disabilities are often reported to have a high prevalence of undetected hearing disorders/loss, but there is no standardized hearing test protocol for this population. The purposes of this study were (1) to examine the hearing status of students with special needs in Taiwan, and (2) to investigate the use of an on-site hearing test protocol that would adequately detect hearing problems in this population and reduce unnecessary referrals for off-site follow-up services. METHODS: A total of 238 students enrolled in two schools for special education and one habilitation center participated in the study. Most students had intellectual disabilities and some also had additional syndromes or disorders. A hearing screening protocol including otoscopy, tympanometry, and distortion product otoacoustic emissions was administered to examine students' outer, middle, and inner ear functions, respectively. Pure tone tests were then administered as an on-site follow-up for those who failed or could not be tested using the screening protocol. RESULTS: Only 32.4% of students passed. When administered alone, the referral rate of otoscopy, tympanometry, and otoacoustic emissions were 38.7%, 46.0%, and 48.5%, respectively. The integration of these subtests revealed 52.1% of students needed follow-up services, 11.8% could not be tested, 2.5% had documented hearing loss, and 1.3% needed to be monitored because of negative middle ear pressure. The inclusion of pure tone audiometry increased the passing rate by 9.9% and provided information on hearing sensitivity for an additional 8.6% of students. CONCLUSION: Hearing assessments and regular hearing screening should be provided as an integral part of health care services for individuals with special needs because of high occurrences of excessive cerumen, middle ear dysfunction, and sensorineural hearing loss. The training of care-givers and teachers of students with special needs is encouraged so that they can help identify hearing problems and reduce the negative impact of hearing disorders and hearing loss. The screening protocol needs to include subtests that examine the status of different parts of their auditory system. The addition of pure tone audiometry as an on-site follow-up tool reduced the rate of off-site referrals and provided more information on hearing sensitivity.