Rapid intracellular pH measurement based on electroporation- surface-enhanced Raman scattering.

In this study, electroporation-surface-enhanced Raman scattering (SERS) was applied to rapidly measure intracellular pH. The generation of a sensitive SERS probe for measuring pH in the range of 6.0-8.0 was accomplished through the conjugation of the pH-sensitive molecule 4-mercaptobenzoic acid (4-MBA) to the surface of gold nanoparticles (Au NPs) through its thiol functional group. This bioprobe was then rapidly introduced into nasopharyngeal carcinoma CNE-1 cells by electroporation, followed by SERS scanning and the fitting of intensity ratios of each detection point's Raman peaks at 1423 cm-1 and 1072 cm-1, to create the pH distribution map of CNE-1 cells. The electroporation-SERS assay introduces pH bioprobes into a living cell in a very short time and disperses the nanoprobe throughout the cytoplasm, ultimately enabling rapid and comprehensive pH analysis of the entire cell. Our work demonstrates the potential of electroporation-SERS for the biochemical analysis of live cells.

Epidermal growth factor potentiates EGFR(Y992/1173)-mediated therapeutic response of triple negative breast cancer cells to cold atmospheric plasma-activated medium.

Cold atmospheric plasma (CAP) holds promise as a cancer-specific treatment that selectively kills various types of malignant cells. We used CAP-activated media (PAM) to utilize a range of the generated short- and long-lived reactive species. Specific antibodies, small molecule inhibitors and CRISPR/Cas9 gene-editing approaches showed an essential role for receptor tyrosine kinases, especially epidermal growth factor (EGF) receptor, in mediating triple negative breast cancer (TNBC) cell responses to PAM. EGF also dramatically enhanced the sensitivity and specificity of PAM against TNBC cells. Site-specific phospho-EGFR analysis, signal transduction inhibitors and reconstitution of EGFR-depleted cells with EGFR-mutants confirmed the role of phospho-tyrosines 992/1173 and phospholipase C gamma signaling in up-regulating levels of reactive oxygen species above the apoptotic threshold. EGF-triggered EGFR activation enhanced the sensitivity and selectivity of PAM effects on TNBC cells. The proposed approach based on the synergy of CAP and EGFR-targeted therapy may provide new opportunities to improve the clinical management of TNBC.

A review on reactive oxygen species (ROS)-inducing nanoparticles activated by uni- or multi-modal dynamic treatment for oncotherapy.

Cancer seriously threatens human health. As compared to normal tissue cells, tumor cells are generally more susceptible to oxidative stress and accumulate higher concentrations of reactive oxygen species (ROS). Accordingly, nanomaterials-based therapies that boost intracellular ROS generation have recently been effective in targeting and eliminating cancer cells by causing programmed death. This review presents a comprehensive analysis of ROS-generation induced by nanoparticles and critically examines the associated therapies which can be categorized as uni-modal (chemodynamic therapy, photodynamic therapy, sonodynamic therapy) and multi-modal (uni-modal therapy + chemotherapy, uni-modal therapy + uni-modal therapy) therapies. Comparison of the relative tumor volume ratio between the experimental and initial tumor volumes shows that multi-modal therapy significantly outperformed other treatments. However, the limitations of multi-modal therapy are in the difficulties of materials preparation and sophisticated operation protocols, thus limiting its applications in clinical practice. As an emerging treatment modality, cold atmospheric plasma (CAP) is a reliable source of ROS, light, and electromagnetic fields that can be used to implement multi-modal treatments in a simple setting. Therefore, the field of tumor precision medicine is expected to increasingly benefit from these promising and rapidly emerging multi-modal therapies based on ROS-generating nanomaterials and reactive media such as CAPs.

Integrated Whole-Exome and Transcriptome Sequencing Indicated Dysregulation of Cholesterol Metabolism in Eyelid Sebaceous Gland Carcinoma.

Purpose: To identify the molecular background of eyelid sebaceous gland carcinomas (SCs), we conducted the integrated whole-exome sequencing and transcriptome sequencing for eyelid SCs in this study. Methods: The genetic alterations were studied by whole-exome sequencing, and the messenger RNA expression was studied using Oxford Nanopore Technologies (ONT) in five paired fresh eyelid SC tissues and adjacent normal tissues. Integrated analysis of exome and transcriptomic information was conducted for filtering candidate driver genes. Protein-protein interaction (PPI) network of filtered candidate genes was analyzed by STRING. The protein expression was verified by immunohistochemistry in 29 eyelid SCs and 17 compared normal sebaceous gland tissues. Results: The average numbers of pathogenic somatic single-nucleotide variants (SNVs) and indels in eyelid SCs were 75 and 28, respectively. Tumor protein p53 (TP53), zinc finger protein 750 (ZNF750), filaggrin 2 (FLG2), valosin-containing protein (VCP), and zinc finger protein 717 (ZNF717) were recurrent mutated genes. A mean of 844 differentially expressed genes (DEGs) were upregulated, and 1401 DEGs were downregulated in SC samples. The intersection of DEG-based pathways and mutation-based pathways was mainly involved in microbial infection and inflammation, immunodeficiency, cancer, lipid metabolism, and the other pathways. The intersection of DEGs and mutated genes consisted of 55 genes, of which 15 genes formed a PPI network with 4 clusters. The PPI cluster composed of scavenger receptor class B member 1 (SCARB1), peroxisome proliferator-activated receptor γ (PPARG), peroxisome proliferator-activated receptor γ coactivator 1α (PPARGC1A) was involved in cholesterol metabolism. The expression of SCARB1 protein was found to be increased, whereas that of PPARG protein was decreased in eyelid SCs compared to that in the normal sebaceous glands. Conclusions: Increased SCARB1 and decreased PPARG indicated that dysregulation of cholesterol metabolism might be involved in carcinogenesis of eyelid SCs. Translational Relevance: The malfunction in cholesterol metabolism might advance our knowledge of the carcinogenesis of eyelid SCs.

Shifted-excitation Raman difference spectroscopy for improving in vivo detection of nasopharyngeal carcinoma.

A strong fluorescence background is one of the common interference factors of Raman spectroscopic analysis in biological tissue. This study developed an endoscopic shifted-excitation Raman difference spectroscopy (SERDS) system for real-time in vivo detection of nasopharyngeal carcinoma (NPC) for the first time. Owing to the use of the SERDS method, the high-quality Raman signals of nasopharyngeal tissue could be well extracted and characterized from the complex raw spectra by removing the fluorescence interference signals. Significant spectral differences relating to proteins, phospholipids, glucose, and DNA were found between 42 NPC and 42 normal tissue sites. Using linear discriminant analysis, the diagnostic accuracy of SERDS for NPC detection was 100%, which was much higher than that of raw Raman spectroscopy (75.0%), showing the great potential of SERDS for improving the accurate in vivo detection of NPC.

Highly Efficient Blood Protein Analysis Using Membrane Purification Technique and Super-Hydrophobic SERS Platform for Precise Screening and Staging of Nasopharyngeal Carcinoma.

Early screening and precise staging are crucial for reducing mortality in patients with nasopharyngeal carcinoma (NPC). This study aimed to assess the performance of blood protein surface-enhanced Raman scattering (SERS) spectroscopy, combined with deep learning, for the precise detection of NPC. A highly efficient protein SERS analysis, based on a membrane purification technique and super-hydrophobic platform, was developed and applied to blood samples from 1164 subjects, including 225 healthy volunteers, 120 stage I, 249 stage II, 291 stage III, and 279 stage IV NPC patients. The proteins were rapidly purified from only 10 µL of blood plasma using the membrane purification technique. Then, the super-hydrophobic platform was prepared to pre-concentrate tiny amounts of proteins by forming a uniform deposition to provide repeatable SERS spectra. A total of 1164 high-quality protein SERS spectra were rapidly collected using a self-developed macro-Raman system. A convolutional neural network-based deep-learning algorithm was used to classify the spectra. An accuracy of 100% was achieved for distinguishing between the healthy and NPC groups, and accuracies of 96%, 96%, 100%, and 100% were found for the differential classification among the four NPC stages. This study demonstrated the great promise of SERS- and deep-learning-based blood protein testing for rapid, non-invasive, and precise screening and staging of NPC.

Multimodal therapy in the management of primary orbital mesenchymal chondrosarcoma.

AIM: To evaluate the ophthalmic manifestations, radiographic features, and prognosis of Chinese patients with primary orbital mesenchymal chondrosarcoma (MCS). METHODS: The study included 6 cases with primary orbital MCS treated at Tianjin Eye Hospital from January 2009 to December 2019. Patitent ophthalmic manifestations, radiographic features, diagnosis, pathology, therapeutic regimens, and prognosis were retrospectively reviewed. RESULTS: Six patitents with primary orbital MCS were identified. The mean age at the first visit was 33y (range, 25-42y). All six patients displayed manifestations of exophthalmos, diplopia, limitation of eye displacement, upper eyelid oedema, decreased visual acuity and ptosis. The mean disease history and range were 5 and 2-8mo, respectively. The tumors were located in the superonasal extraconal compartment (2/6, 33.3%), intraconal compartment (2/6, 33.3%), and bitemporal extraconal compartment (2/6, 33.3%), respectively. Radiographic features were a well-defined, orbital mass with calcification and ossification on computed tomography (CT), and marked heterogenous enhancement on dynamic magnetic resonance imaging (MRI). Five patients were treated with tumor resection and one patient received orbital exenteration. Five patients in the cohort received postoperative radiation therapy, two patients received chemotherapy, and one patient did not receive postoperative adjuvant therapy because he refused. The histopathologic classification revealed a tumour composed of a mixture of mature chondroid tissue surrounded by small, round, and undifferentiated mesenchymal cells. Immunohistochemistry revealed Bcl-2, vimetin, CD99, and S-100 were expressed were expressed. After surgeries, two patients have developed a local recurrence. The median recurrence time of 58mo (52-64mo). One patient had distant recurrence included the lungs occurred 52mo after the initial surgery. CONCLUSION: The possibilty of orbital MCS need to be considered when a painless, slowly growing orbital mass with calcification and ossification. From our experience, trimodality treatment of radiation therapy, chemotherapy and surgery maybe the best option. Orbital MCS has a high tendency for late recurrence, regular long-term follow-up after complete excision is mandatory.

Modified reduced-volume intensity-modulated radiation therapy in non-metastatic nasopharyngeal carcinoma: A prospective observation series.

BACKGROUND AND PURPOSE: Although the efficacy of "reduced-volume intensity-modulated radiation therapy (IMRT)" in nasopharyngeal carcinoma (NPC) has been confirmed, two issues regarding the necessity of clinical target volume 1(CTV1) delineation and the optimal margin of CTV2 remained undetermined. The current series, utilized de-intensification technique that omitted the contouring of CTV1 and narrowed the margin of CTV2 from 10 mm to 8 mm, namely "modified reduced-volume IMRT" was initiated to evaluate the efficacy and feasibility of this renew technique in a prospective series. PATIENTS AND MATERIALS: Dosimetric analysis was performed in 40 non-metastatic NPC cases to evaluate whether our modification is feasible. Then this de-intensification technique was applied in non-metastatic NPC patients treated in our attending group since late 2014. Survival outcomes focused on local recurrence-free survival (LRFS) and local failure pattern were analyzed. RESULTS: Preliminary dosimetric evaluation of "modified reduced-volume IMRT" showed that the 60 Gy isodose curve generated naturally by this technique could well wrap the target area of CTV1. Subsequent observation series, which included a total of 471 patients and had a median follow-up time of 46.2 months(range,3.7-70.8 months), reported that 4-year estimated LRFS, regional recurrence-free survival (RRFS), distant metastasis-free survival (DMFS) and overall survival (OS) were 96.6%, 97.7%, 87.7% and 92.4%, respectively. All local recurrence lesions occurred within 95% isodose lines and were considered in-field failures. CONCLUSIONS: Our de-intensification technique "modified reduced-volume IMRT" was feasible and did not compromise therapeutic efficacy, well-designed multicenter prospective trials are needed for further research.

Primary orbital extraskeletal osteosarcoma and review of literature.

BACKGROUND: Extraskeletal osteosarcoma is a malignant tumour composed of an osteoid and/or cartilaginous matrix; it arises in soft tissues without connection to the skeleton, and to our knowledge, this type of tumour is extremely rare. CASE PRESENTATION: The present study reports a 57-year-old man with primary orbital extraskeletal osteosarcoma who presented with a history of painful swelling in the left orbit that had occurred for 11 months. Imaging of the orbit showed an atypical, well-defined heterogeneous mass attached to the posterior globe of the left orbit. The patient underwent an anterior orbitotomy and complete excision of the tumour. The mass was originated from neither the globe nor the bony orbital wall but from the soft tissue. Histopathology demonstrated an extraskeletal osteosarcoma. After 13 months of follow-up, there was apparent recurrence of the tumour. The medical history showed no complaints of previous trauma or radiotherapy. CONCLUSIONS: ESOS is a highly malignant tumour. Immunosuppression, trauma and adjuvant radiotherapy are possible predisposing factors in the development of this tumour. Prompt recognition and thorough treatment are essential for preventing orbital lesions and presence of metastasis from other organs.

Severe Marginal Ulcerative Keratitis Induced by an Insect Foreign Body in the Cornea.

PURPOSE: To report an uncommon case in which an insect foreign body (FB) in the cornea induced severe marginal ulcerative keratitis. METHODS: The insect body was removed during an eye examination. A shaped deep lamellar keratoplasty was then performed on the ulcerous corneal tissue and pseudopterygium. The insect FB was identified as a member of the nontoxic Diptera insect order, belonging to either the Chloropidae or Sphaeroceridae family. RESULTS: The patient recovered well after the keratoplasty, and her corneal graft was clear with significantly improved visual acuity after 2 months. CONCLUSIONS: Long-time retention of a nontoxic insect FB in the cornea could induce severe noninfectious keratitis. Individually designed deep lamellar keratoplasty successfully restored the transparency of the cornea and visual function.

Rapid and label-free urine test based on surface-enhanced Raman spectroscopy for the non-invasive detection of colorectal cancer at different stages.

The concept of being able to urinate in a cup and screen for colorectal cancer (CRC) is fascinating to the public at large. Here, a simple and label-free urine test based on surface-enhanced Raman spectroscopy (SERS) was employed for CRC detection. Significant spectral differences among normal, stages I-II, and stages III-IV CRC urines were observed. Using discriminant function analysis, the diagnostic sensitivities of 95.8%, 80.9%, and 84.3% for classification of normal, stages I-II, and stages III-IV CRC were achieved in training model, indicating the great promise of urine SERS as a rapid, convenient and noninvasive method for CRC staging detection.

What is the impact of intravitreal injection of conbercept on neovascular glaucoma patients: a prospective, interventional case series study.

BACKGROUND: The aim of the present study was to evaluate the efficacy and safety of intravitreal conbercept combined with trabeculectomy and panretinal photocoagulation for neovascular glaucoma (NVG). METHODS: Fifty patients (54 eyes) with NVG were included in this prospective study. Fifty-two eyes initially underwent intravitreal conbercept (0.5 mg/0.05 ml) treatment followed by trabeculectomy and panretinal photocoagulation. Preoperative and postoperative best-corrected visual acuity (BCVA), intraocular pressure (IOP), the number of antiglaucoma medications, and surgical complications were recorded. The levels of VEGF-A, TGF-ß1 and PLGF in aqueous humour samples collected during surgery were measured by enzyme-linked immunosorbent assay (ELISA). Light microscopy and transmission electron microscopy were used to observe the surgically excised trabecular tissue; enucleation was performed in 2 eyes, and light microscopy was used as the histopathological control. RESULTS: The follow-up period after trabeculectomy was 1 year. Of the 52 eyes, 39 completed 1 year of follow-up, and 13 were lost to follow-up. Recurrence of iris neovascularization was observed in 5 eyes, 9 had hyphema, 16 had filter-bled scarring, and no eye had complications attributable to the drug. The mean IOP was reduced from 48.1 ± 14.2 to 23.2 ± 8.7 mmHg, and the mean number of antiglaucoma medications used decreased from 3.0 (3.0, 4.0) to 1.0 (0.0, 1.0) after 1 year (both P < 0.05). The complete success rate was 76.9, 76.9, 71.0, 51.6, and 32.3% at 1 week, 1 month, 3 months, 6 months and 12 months, respectively, when the cut-off IOP was 18 mmHg. After patients underwent intravitreal injection, the concentrations of VEGF-A and TGF-ß1 in the aqueous humour in NVG patients decreased from 168.8 ± 13.4 and 159.6 ± 15.4 pg/ml to 160.2 ± 7.6 and 151.9 ± 2.3 pg/ml, respectively (both P < 0.05). Light microscopy revealed neovascularization regression in the iris in specimens treated with intravitreal conbercept. Electron microscopy revealed trabecular endothelial cell degeneration in the conbercept-treated specimens. CONCLUSIONS: Our initial findings suggest that intravitreal conbercept is an effective treatment for managing NVG that has fewer short-term postoperative complications. TRIAL REGISTRATION: Current Controlled Trials ChiCTR1800019918 , 8 December 2018, retrospectively registered.

A novel urine analysis technique combining affinity chromatography with Au nanoparticle based surface enhanced Raman spectroscopy for potential applications in non-invasive cancer screening.

Modified nucleoside in urine samples is one of the most common biomarkers for cancer screening. Therefore, we developed a novel detection method for modified nucleoside detection in human urine. In this work, the modified nucleoside from real cancer patient's urine samples was first separated and purified using the affinity chromatography (AC) technology relying on its specific adsorption capacity. Then, surface-enhanced Raman spectroscopy (SERS) technology with the capability of single molecular detection was used to sensitively characterize the biomolecular features of modified nucleoside. A total of 141 high-quality SERS spectra of urinary modified nucleoside can be obtained from 50 gastric cancer patients and 43 breast cancer patients, as well as 48 healthy volunteers. Using principal component analysis combined with linear discriminant analysis (PCA-LDA), the diagnostic sensitivities for identifying gastric cancer vs normal, breast cancer vs normal, gastric cancer vs breast cancer were 84.0%, 76.7% and 82.0%, respectively, and the corresponding diagnostic specificities for each combination were 95.8%, 87.5% and 90.7%, respectively. These results show that this novel method based on urinary modified nucleoside detection combining AC and SERS technologies holds promising potential for developing a specific, non-invasive and label-free tool for cancer screening.

Positive relationship between number of negative lymph nodes and duration of gallbladder cancer cause-specific survival after surgery.

BACKGROUND: Although the prognostic implications of negative lymph nodes (NLNs) has been reported for a variety of tumors, little information has been published about the NLNs in gallbladder cancer (GBC). PATIENTS AND METHODS: In this study, clinicopathological characteristics and survival times of patients who had undergone surgery for GBC were collected from the Surveillance, Epidemiology, and End Results Program-registered TNM stage database and analyzed. Univariate and multivariate Cox proportional hazards models were used to identify the predictors of survival. RESULTS: It was found that a cutoff of one to two NLNs is optimal when assessing the association with survival, survival rates being consistently better with two or more NLNs than with fewer than two. This optimal cutoff value of 2 was identified as an independent prognostic factor by univariate and multivariate analyses (all P<0.001). Specifically, patients with two or more NLNs had better 5-year gallbladder cancer cause-specific survival than those with fewer than NLNs examined for stage I/II, stage III/IV, and all TNM stages (all P<0.001). CONCLUSION: Our findings indicate that the number of NLNs is an independent prognostic factor after GBC surgery, and, together with the number of positive lymph nodes, this will provide better prognostic information than the number of positive lymph nodes alone.

A retrospective study of 2228 cases with eyelid tumors.

AIM: To describe the histopathologic and clinical features of eyelid tumor cases from Tianjin Eye Hospital during 2002 to 2015. METHODS: In this retrospective study, a total of 2228 cases of eyelid tumors with pathologic diagnoses were enrolled. The eyelid tumors were classified into three groups according to tumor origin: epidermal, adnexal and miscellaneous, including melanocytic, neural and vascular lesions. Inflammatory tumor-like lesions were excluded. The clinical characteristics of the eyelid tumors were analyzed, including age, gender and lesion location. RESULTS: Most eyelid tumors were epidermal in origin (1080, 48.5%), followed by miscellaneous (885, 39.7%) and adnexal tumors (263, 11.8%). Among all the tumors, 292 (13.1%) were malignant lesions, 1910 (85.7%) benign and 26 (1.1%) premalignant lesions. Most malignant tumors originated from epidermal cells (60.0%), followed by adnexal cells (34.6%). The most common malignant tumors were basal cell carcinomas (56.5%) followed by sebaceous carcinoma (34.6%), squamous cell carcinomas (3.8%) and lymphoma/plasmocytoma (1.7%). The benign and premalignant eyelid lesions mostly originated from epidermal cells (46.4%) followed by miscellaneous cell sources (45.2%), including melanocytic nevus (33.8%), seborrheic keratosis (13.7%), squamous cell papilloma (13.0%) and epidermal cysts (11.5%). CONCLUSION: Eyelid tumors are mostly epithelial in origin. Benign tumors are significantly more common than malignant tumors with an obvious female predominance, and the most frequent malignant tumor are basal cell carcinoma, sebaceous carcinoma and squamous cell carcinomas. The tumor clinical features varied among the different subtypes.

Label free hepatitis B detection based on serum derivative surface enhanced Raman spectroscopy combined with multivariate analysis.

Surface-enhanced Raman spectroscopy (SERS) was developed here for the non-invasive detection of the hepatitis B virus (HBV). Chronic hepatitis B virus (HBV) infection is a primary health problem in the world and may further develop into cirrhosis and hepatocellular carcinoma (HCC). SERS measurement was applied to two groups of serum samples. One group included 93 HBV patients and the other group included 94 healthy volunteers as control subjects. Tentative assignments of the Raman bands in the measured SERS spectra have shown the difference of the serum SERS spectra between HBV patients and healthy volunteers. The differences indicated an increase in the relative amounts of L-arginine, Saccharide band (overlaps with acyl band), phenylalanine and tyrosine, together with a decrease in the percentage of nucleic acid, valine and hypoxanthine in the serum of HBV patients compared with those of healthy volunteers. For better analysis of the spectral data, the first-order derivation was applied to the SERS data. Furthermore, principal components analysis (PCA), combined with linear discriminant analysis (LDA), were employed to distinguish HBV patients from healthy volunteers and to realize the diagnostic sensitivity of 78.5% and 91.4%, and specificity of 75% and 83% for SERS and the first order derivative SERS spectrum, respectively. These results suggest that derivative analysis could be an effective method to improve the classification of SERS spectra belonging to different groups. This exploratory work demonstrated that first-order derivative serum SERS spectrum combined with PCA-LDA has great potential for improving the detection of HBV.

Future Trend Forecast by Empirical Wavelet Transform and Autoregressive Moving Average.

In engineering and technical fields, a large number of sensors are applied to monitor a complex system. A special class of signals are often captured by those sensors. Although they often have indirect or indistinct relationships among them, they simultaneously reflect the operating states of the whole system. Using these signals, the field engineers can evaluate the operational states, even predict future behaviors of the monitored system. A novel method of future operational trend forecast of a complex system is proposed in this paper. It is based on empirical wavelet transform (EWT) and autoregressive moving average (ARMA) techniques. Firstly, empirical wavelet transform is used to extract the significant mode from each recorded signal, which reflects one aspect of the operating system. Secondly, the system states are represented by the indicator function which are obtained from those normalized and weighted significant modes. Finally, the future trend is forecast by the parametric model of ARMA. The effectiveness and practicality of the proposed method are verified by a set of numerical experiments.

Application of a near-infrared laser tweezers Raman spectroscopy system for label-free analysis and differentiation of diabetic red blood cells.

A home-made near-infrared laser tweezers Raman spectroscopy (LTRS) system was applied to detect hemoglobin variation in red blood cells (RBCs) from diabetes without exogenous labeling. Results showed significant spectral differences existed between the diabetic and normal RBCs, including the peaks dominated by protein components (e.g. 1003 cm-1) and heme groups (e.g. 753 cm-1) in RBCs, and accurate classification results for diabetes detection were obtained by linear discriminant analysis with 100% sensitivity (i.e. no false negatives in the study). This work indicated the great promise of LTRS as a label-free RBC analytical tool for improving the accurate detection of type II diabetes.

Assessment of p16 expression and HPV infection in adenoid cystic carcinoma of the lacrimal gland.

Purpose: Adenoid cystic carcinoma (ACC) in the lacrimal gland is a rare malignancy. P16 is encoded by the CDKN2A gene, which is recognized as a tumor suppressor due to its inactivation in many types of tumors. However, p16 overexpression is also linked to adverse tumor parameters. These contradictory observations have also been confirmed in ACCs in the salivary glands. Furthermore, evidence of human papilloma virus (HPV) infection is found in a proportion of ACCs in the salivary glands. P16 is often overexpressed in HPV-related squamous cell carcinoma in parallel. To our knowledge, the role of p16 and HPV in ACCs in the lacrimal gland is still unknown. Methods: Twenty-one ACCs in the lacrimal gland and ten matched healthy lacrimal glands were studied. P16 was detected with immunohistochemistry (IHC), and HPV was detected with in situ hybridization (ISH) and PCR in all cases. Other cell cycle proteins were also detected with IHC, including cyclin D1 and Ki67. The methylation status of the p16 promoter was detected with methylation-specific PCR (MSP) to further investigate the regulation of p16 expression. Results: The expression rates of p16 (47.6%, 10/21), cyclin D1 (100%, 21/21), and Ki67 (52.4%, 11/21) were increased in ACCs compared to healthy lacrimal glands (negative). The results showed p16 expression was limited to the inner ductal epithelial cells in the majority of the tubular and cribriform patterns. In solid ACCs, p16 was uniformly positive. HPV was negative in all 21 cases with ISH and PCR. P16 overexpression was associated with cyclin D1 overexpression (p=0.013). Only 13 cases were tested successfully with MSP. The expression rate of p16 methylation was 23.1% (3/13) of the ACCs. Compared with primary ACCs, recurrent ACCs showed higher p16, cyclin D1, and Ki67 expression (p=0.011, p=0.026, p=0.049, respectively). Conclusions: In summary, p16 overexpression was cell-type dependent in ACCs in the lacrimal gland, while HPV infection was negative. P16 overexpression was unrelated to HPV infection. The mechanism of p16 overexpression needs to be further investigated in ACCs in the lacrimal gland.

Expression of Peroxiredoxin 2 and Vascular Endothelial Growth Factor Receptor 2 in Pterygium.

PURPOSE: The expression of peroxiredoxin 2 and vascular endothelial growth factor receptor 2 (VEGFR2) was detected in pterygium to investigate whether they are involved in the pathogenesis or recurrence of pterygium and to evaluate the association between peroxiredoxin 2 and VEGFR2 in pterygium. METHODS: Ten normal bulbar conjunctivae, 35 primary pterygia, and 35 recurrent pterygia were obtained. Formalin-fixed, paraffin-wax-embedded tissues were analyzed by immunohistochemistry with peroxiredoxin 2 and VEGFR2 antibodies. RESULTS: There was no statistical difference between primary pterygia and recurrent pterygia in terms of age and sex (P = 0.685; P = 0.811). The expression rate of peroxiredoxin 2 (94.3%, 66/70) and VEGFR2 (61.4%, 43/70) was increased in pterygia compared with normal conjunctivae (negative). The expression of peroxiredoxin 2 in recurrent pterygia (negative 0, weak 0, moderate 27, strong 8) was higher than that in primary pterygia (negative 6, weak 16, moderate 13, strong 0) (P < 0.001). The expression of VEGFR2 in recurrent pterygia (negative 4, weak 5, moderate 12, strong 4) was higher than that in primary pterygia (negative 23, weak 10, moderate 1, strong 1) (P < 0.001). The expression of peroxiredoxin 2 was consistent with that of VEGFR2 in pterygium (r = 0.348, P = 0.006). CONCLUSIONS: Overexpression of peroxiredoxin 2 and VEGFR2 in pterygium might be involved in the pathogenesis or recurrence of pterygium. The increase of VEGFR2 might be related to the increase of peroxiredoxin 2 in response to excessive reactive oxygen species from ultraviolet exposure.