Randomized Controlled Trial Investigating Perioperative Immunonutrition for Patients Undergoing Cytoreductive Surgery (CRS) and Hyperthermic Intraperitoneal Chemotherapy (HIPEC).

BACKGROUND: Immunonutrition has been shown to reduce hospital stay and postoperative morbidity in patients undergoing gastrointestinal, and head and neck surgery. However, its use has not been demonstrated in patients undergoing cytoreductive surgery and hyperthermic intraperitoneal chemotherapy (CRS-HIPEC). This study aims to determine the effectiveness of perioperative immunonutrition on patients undergoing CRS-HIPEC in reducing length of hospitalization and postoperative complications. PATIENTS AND METHODS: From April 2017 to December 2018, patients undergoing CRS-HIPEC for peritoneal metastases in a single center were enrolled in a randomized controlled trial. Patients with evidence of intestinal obstruction or with diabetes mellitus were excluded. Patients were randomly assigned in a 1:1 fashion to receive perioperative oral immunonutrition or standard nutritional feeds. Length of hospital stay and rates of wound infection and complications were recorded and compared between the two groups in an intention-to-treat manner. RESULTS: A total of 62 patients were recruited and randomized into two groups. Compliance to nutritional feeds in the preoperative period was significantly higher in the standard nutrition group (95.2% versus 75.4%, p = 0.004). There was no difference in postoperative compliance rates. Length of hospital stay and rates of wound infection and postoperative complications were higher in the standard nutrition group when compared with patients on immunonutrition (15.5 versus 11.1 days, p = 0.186; 19% versus 9.7%, p = 0.473; 16% versus 9.7%, p = 0.653; respectively). CONCLUSIONS: Patients undergoing CRS-HIPEC who received perioperative immunonutrition had shorter hospitalization and less wound infections and postoperative complications, although the differences with the standard nutrition group were not statistically significant. Potential benefits of perioperative immunonutrition need to be further evaluated in larger studies.

Usage of glaucoma-specific patient-reported outcome measures (PROMs) in the Singapore context: a qualitative scoping exercise.

BACKGROUND: Despite the increasing emphasis on the role of glaucoma-specific patient-reported outcome measures (PROMs) as relevant outcome measures for the impact of glaucoma and its intervention on patients' daily lives, the feasibility of implementing PROMs in the routine clinical setting in Singapore remains undefined. We aim to evaluate the comprehensibility, acceptability, and relevance of four glaucoma-specific PROMs at healthcare professionals' and patients' level in a Singapore context. METHODS: Sixteen ophthalmic healthcare professionals and 24 glaucoma patients, with average age 60 years (SD = 15), were invited from a tertiary hospital in Singapore. Semi-structured interviews were conducted to explore participants' perceptions on the content and administration of four glaucoma-specific PROMs - the Glaucoma Quality of Life-15, Glaucoma Symptom Identifier, Independent Mobility Questionnaire and Treatment Satisfaction Survey of Intra-ocular Pressure. Semi-structured interviews were hand transcribed, and analysed thematically. Each participant filled out a feasibility survey at the end of interview. RESULTS: 79% of glaucoma patients and 94% of glaucoma healthcare professionals felt selected PROMs relevant to patients. 63% of glaucoma patients and 50% of healthcare professionals felt that selected PROMs were sufficiently comprehensive for clinical use. 46% of glaucoma patients and 56% of healthcare professionals felt selected PROMs were user-friendly. CONCLUSIONS: Using PROMs in the Singapore clinical setting receives promising support from both healthcare professionals and patients. The identified potential barriers tailored to Singapore clinical setting will help successful implementation of PROMs into routine clinical care.

Rational Molecular Design of Potent PLK1 PBD Domain-binding Phosphopeptides Using Preferential Amino Acid Building Blocks.

Polo-like kinase 1 (PLK1) is an important regulator in diverse aspects of the cell cycle and proliferation. The protein has a highly conserved polo-box domain (PBD) present in C-terminal noncatalytic region, which exhibits a relatively broad sequence specificity in recognizing and binding phosphorylated substrates to control substrate phosphorylation by the kinase. In order to elucidate the structural basis, thermodynamic property, and biological implication underlying PBD-substrate recognition and association, a systematic amino acid preference profile of phosphopeptide interaction with PLK1 PBD domain was established via virtual mutagenesis analysis and mutation energy calculation, from which the contribution of different amino acids at each residue position of two reference phosphopeptides to domain-peptide binding was characterized comprehensively and quantitatively. With the profile, we are able to determine the favorable, neutral, and unfavorable amino acid types for each position of PBD-binding phosphopeptides, and we also explored the molecular origin of the broad sequence specificity in PBD-substrate recognition. To practice computational findings, the profile was further employed to guide rational design of potent PBD binders; three 6-mer phosphopeptides (i.e., IQSpSPC, LQSpTPF, and LNSpTPT) were successfully developed, which can efficiently target PBD domain with high affinity (Kd = 5.7 ± 1.1, 0.75 ± 0.18, and 7.2 ± 2.6 µm, resp.) as measured by a fluorescence anisotropy assay. The complex structure of PLK1 PBD domain with a newly designed, potent phosphopeptide LQSpTPF as well as diverse noncovalent chemical forces, such as H-bonds and hydrophobic interactions at the complex interface, were examined in detail to reveal the molecular mechanism of high affinity and stability of the complex system.

Moesin as a key cytoskeleton regulator in corneal fibrosis.

PURPOSE: : Corneal fibrosis is the third leading cause of blindness worldwide. α-Smooth muscle actin (SMA), a marker of fibrosis, is closely regulated through an intermediate group of submembrane molecules - cytoskeleton regulators. The purpose of this study was to elucidate the role of specific cytoskeleton regulators in a mouse model of corneal fibrosis. METHODS: : A mouse model of corneal fibrosis was developed using anterior keratectomy (AK) and the topical application of transforming growth factor (TGF)-ß1 (1 µg/ml). The RT² Profiler™ PCR Array for cytoskeleton regulators was used to assay changes in levels of specific members of this class of proteins. Moesin siRNA was delivered into the corneal stroma by iontophoresis in vivo. Transformation of the corneal keratocyte-to-myofibroblast in corneal fibrosis, as defined by the expression of α-SMA, was determined by Western blot. RESULTS: : After AK and topical application of TGF-ß1, moesin was the most highly upregulated gene among 84 cytoskeleton regulator genes; iontophoresing moesin siRNA into the corneal stroma reduced the expression of α-SMA to 0.22-, 0.52-, and 0.31-fold of control at postoperative (PO) day 1, 3, and 5, respectively; also, upregulation of phospho-Smad 2 induced by TGF-ß1 was reduced by moesin siRNA to 0.59-, 0.56-, and 0.31-fold of control and expression of phospho-Smad 3 was reduced to 0.58-, 0.53-, and 0.47-fold of control at the same PO days. CONCLUSIONS: : Moesin may be a potential drug target for inhibiting corneal fibrosis, and the details of moesin-related signaling pathways would be critical for understanding corneal fibrosis.

Incidence and outcomes of delayed presentation and surgery in peritoneal surface malignancies.

Background: Peritoneal surface malignancies (PSM) present insidiously and often pose diagnostic challenges. There is a paucity of literature quantifying the frequency and extent of therapeutic delays in PSM and its impact on oncological outcomes. Methods: A review of a prospectively maintained registry of PSM patients undergoing Cytoreductive Surgery and Hyperthermic Intra-peritoneal Chemotherapy (CRS-HIPEC) was conducted. Causes for treatment delays were identified. We evaluate the impact of delayed presentation and treatment delays on oncological outcomes using Cox proportional hazards models. Results: 319 patients underwent CRS-HIPEC over a 6-years duration. 58 patients were eventually included in this study. Mean duration between symptom onset and CRS-HIPEC was 186.0 ± 37.1 days (range 18-1494 days) and mean duration of between patient-reported symptom onset and initial presentation was 56.7 ± 16.8 days. Delayed presentation (> 60 days between symptom onset and presentation) was seen in 20.7% (n=12) of patients and 50.0% (n=29) experienced a significant treatment delay of > 90 days between 1st presentation and CRS-HIPEC. Common causes for treatment delays were healthcare provider-related i.e. delayed or inappropriate referrals (43.1%) and delayed presentation to care (31.0%). Delayed presentation was a significantly associated with poorer disease free survival (DFS) (HR 4.67, 95% CI 1.11-19.69, p=0.036). Conclusion: Delayed presentation and treatment delays are common and may have an impact on oncological outcomes. There is an urgent need to improve patient education and streamline healthcare delivery processes in the management of PSM.

A polycationic antimicrobial and biocompatible hydrogel with microbe membrane suctioning ability.

Despite advanced sterilization and aseptic techniques, infections associated with medical implants have not been eradicated. Most present coatings cannot simultaneously fulfil the requirements of antibacterial and antifungal activity as well as biocompatibility and reusability. Here, we report an antimicrobial hydrogel based on dimethyldecylammonium chitosan (with high quaternization)-graft-poly(ethylene glycol) methacrylate (DMDC-Q-g-EM) and poly(ethylene glycol) diacrylate, which has excellent antimicrobial efficacy against Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus and Fusarium solani. The proposed mechanism of the antimicrobial activity of the polycationic hydrogel is by attraction of sections of anionic microbial membrane into the internal nanopores of the hydrogel, like an 'anion sponge', leading to microbial membrane disruption and then microbe death. We have also demonstrated a thin uniform adherent coating of the hydrogel by simple ultraviolet immobilization. An animal study shows that DMDC-Q-g-EM hydrogel coating is biocompatible with rabbit conjunctiva and has no toxicity to the epithelial cells or the underlying stroma.

Can baseline quality of life scores predict for morbidity and survival after CRS and HIPEC: a prospective study of 151 patients.

Objectives: Various studies have shown that good quality of life (QoL) can be achieved after cytoreductive surgery (CRS) and hyperthermic intraperitoneal chemotherapy (HIPEC). There is prognostic value of baseline QoL in post-operative outcome in Western setting. Our prospective study aims to validate these observations and elucidate clinical factors that predict poorer QoL in Asian peritoneal carcinomatosis patients. Methods: European Organization for Research and Treatment of Cancer Core Quality of Life Questionnaire was administered to patients before CRS and HIPEC and thereafter at 3, 6 and 12 months. Results: A total of 151 patients underwent 155 surgeries. Four hundred and seventy two questionnaires were completed. Median disease-free survival (DFS) was 16.5 months. Three year DFS and overall survival (OS) were 24.0% and 73.0% respectively. Post-operative global health status significantly increased at 3, 6 and 12 months. The decreases in functional scales recovered to baseline by 1-year post-surgery. Peritoneal carcinomatosis index (PCI), presence of stoma, peritonectomy duration, death within one year, post-operative complication and length of SICU stay negatively influenced QoL. Complication rates were higher in patients with lower global health status, physical and role functioning scores and higher symptom summary scores at baseline. Lower social functioning score, and higher pain, dyspnoea and symptom summary scores at baseline were significantly associated with poorer OS. Conclusions: Various clinical factors can help us predict a patient's QoL after surgery. Several baseline factors were also able to predict morbidity and survival. Going forward, we can use these factors to help us better select patients who will have a greater benefit from CRS and HIPEC.

Trajectories of Patient-Reported Outcomes After Palliative Gastrointestinal Surgery in Advanced Cancer: Is Good Quality of Life Sustainable?

To evaluate the trajectories and sustainability of health-related quality of life (HRQoL) outcomes after palliative gastrointestinal (GI) surgery and perioperative factors associated with HRQoL improvement postsurgery. Background: Palliative patients face a wide range of physical, emotional, social, and functional challenges. In evaluating the efficacy of palliative surgical interventions, a major pitfall of traditional surgical outcome measures is that they fall short of measuring outcomes that are meaningful to patients during end-of-life. HRQoL tools may provide a more comprehensive assessment of the true value and impact of palliative surgery. Methods: We prospectively recruit advanced cancer patients undergoing palliative GI surgery. The Functional Assessment of Cancer Therapy-General (FACT-G) questionnaire was administered before and at regular intervals after surgery. HRQoL improvement was defined as ≥4-points increment in FACT-G total score over baseline. Duration of sustained HRQoL improvement above this threshold and factors associated with varying extents of HRQoL change were evaluated. Results: Of the 65 patients, intestinal obstruction was the most common indication for surgery (70.8%). The mean baseline FACT-G total score was 70.7 (95% CI: 66.3-75.1). Forty-six (70.8%) patients experienced HRQoL improvement after surgery. This HRQoL improvement was sustained over a median duration of 3.5 months and was driven mainly by improvements in patients' physical and emotional well-being. Albumin was significantly associated with the extent of HRQoL improvements (P = 0.043). Conclusion: A clinically significant and sustained improvement in HRQoL was observed after palliative GI surgery. Patients with higher preoperative albumin levels were more likely to experience HRQoL improvements.

Generating patient-derived ascites-dependent xenograft mouse models of peritoneal carcinomatosis.

Clinically relevant animal models are crucial for effective development of therapeutics for peritoneal carcinomatosis (PC). This protocol describes the generation of patient-derived ascites-dependent xenograft (PDADX) models from the cellular component of ascites. The use of routine intraperitoneal injection of the fluid component of ascites is analogous to the biological events occurring intra-abdominally in patients with PC. By serving as a proxy, PDADX models represent a valuable tool for preclinical testing of new therapeutics for PC. For complete details on the use and execution of this protocol, please refer to Hendrikson et al. (2022).

Prospective Comparison of the Performance of MRI Versus CT in the Detection and Evaluation of Peritoneal Surface Malignancies.

Background: The performance of MRI versus CT in the detection and evaluation of peritoneal surface malignancies (PSM) remains unclear in the current literature. Our study is the first prospective study in an Asian center comparing the two imaging modalities, validated against intra-operative findings. Methods: A total of 36 patients with PSM eligible for CRS-HIPEC underwent both MRI and CT scans up to 6 weeks before the operation. The scans were assessed for the presence and distribution of PSM and scored using the peritoneal cancer index (PCI), which were compared against PCI determined at surgery. Results: Both MRI and CT were 100% sensitive and specific in detecting the overall presence of PSM. Across all peritoneal regions, the sensitivity and specificity for PSM detection was 49.1% and 93.0% for MRI, compared to 47.8% and 95.1% for CT (p = 0.76). MRI was more sensitive than CT for small bowel disease, although the difference did not reach statistical significance. Comparing PCI on imaging with intra-operative PCI, the mean difference was found to be −3.4 ± 5.4 (p < 0.01) for MRI, and −3.9 ± 4.1 (p < 0.01) for CT. The correlation between imaging and intra-operative PCI was poor, with a concordance coefficient of 0.76 and 0.79 for MRI and CT, respectively. Within individual peritoneal regions, there was also poor agreement between imaging and intra-operative PCI for both modalities, other than in regions 1 and 3. Conclusion: MRI and CT are comparable in the detection and evaluation of PSM. While sensitive in the overall detection of PSM, they are likely to underestimate the true disease burden.

Epithelial microfilament regulators show regional distribution in mouse conjunctiva.

PURPOSE: The conjunctival epithelium is a continuous sheet of cells with regional characteristics that appear to be similar. This study was designed to investigate the distribution and levels of expression of a subset of microfilament regulators in the forniceal, palpebral, and bulbar conjunctival epithelia. METHODS: Balb/C mice were used. The localizations of paxillin, focal adhesion kinase, vinculin, talin1, cofilin, profilin, gelsolin, integrin ß1, and integrin α6 were studied with the use of cross-sectional immunofluorescent staining. For a detailed cellular analysis, positioning and ablation with the laser microbeam (PALM) Combi System was used to obtain forniceal, bulbar, and palpebral conjunctival epithelia for expression comparison with the use of western blot analysis and quantitative real-time polymerase chain reaction. RESULTS: Immunostaining showed that focal adhesion kinase, cofilin, profilin, gelsolin, talin1, and vinculin were expressed in all layers of the forniceal, palpebral, and bulbar conjunctival epithelia. Paxillin, integrin ß1, and α6 was found to be located in the basal cell layer in all three of these areas. Quantitative real-time polymerase chain reaction showed that the transcript levels of these microfilament regulators in the forniceal conjunctivae were higher than those levels found in the bulbar and palpebral conjunctivae. Western blot analysis confirmed the differential expression levels of these microfilament regulators in the forniceal, bulbar, and palpebral conjunctivae. CONCLUSIONS: Differences in the levels of microfilament regulators in the forniceal, bulbar, and palpebral conjunctivae suggest different modes of interaction with their microenvironment and within cell layers.

[Metabonomic study of the urine of rat acute gouty arthritis model].

UNLABELLED: [ OBJECTIVE: To analysis the metabonome characteristics of the urine of MSU crystal-induced acute gouty arthritis rats by the method of metabolomics. METHODS: Based on the method of metabolomics, which applies LC/MS as data acquisition platform, incorporating with the means of stoechiometry such as principal component analysis, we analyzed the metabonome difference between the urine of acute gouty arthritis rats and that of normal rats. RESULTS: Compare with control group, the metabolism status of acute gouty arthritis model group deviated. After that, with the time lapsed, it retrieved gradually to the incipient metabolism status. The variation of metabolism locus of rats measured by the methods of metabolomics properly reflects the genesis, development, and recuperation course of acute gouty arthritis. CONCLUSION: The whole metabolism status of rat model is able to be presented finely with the method of metabolomics. The metabolomics study could offer a satisfactory research approach to acute gouty arthritis.

Fluorescein Labeled Leukocytes for in vivo Imaging of Retinal Vascular Inflammation and Infiltrating Leukocytes in Laser-Induced Choroidal Neovascularization Model.

Purpose: To study the effect of anti-VEGF treatment on retinal inflammation in a laser-induced CNV rodent model.Methods: Leukocytes labeled with 1% sodium fluorescein were injected into the laser-induced CNV (wild type C57BL/6) mice at days 4 (baseline), 7, 14, and 19. At baseline intravitreally 3 mice received 1× PBS, and 3 mice received anti-VEGF. FFA, OCT, and SLO were performed at each time point to assess the CNV pathophysiology and inflammatory response.Results: Fluorescein leakage, SRF, and leukocyte infiltration were observed at baseline in both the groups before injection. From days 7 to 19, leukocyte infiltration and SRF were noted in the 1× PBS group, but limited or no SRF and leukocyte infiltration was observed in the anti-VEGF group.Conclusions: Leukocyte infiltration was established as an in vivo imaging inflammatory marker and along with FFA and OCT showed response to anti-VEGF therapy in laser-induced CNV model.

Clinical evidence for association of neoadjuvant chemotherapy or chemoradiotherapy with efficacy and safety in patients with resectable esophageal carcinoma (NewEC study).

BACKGROUND: The efficacy and safety of neoadjuvant treatment over surgery alone and that of neoadjuvant chemoradiotherapy (NCRT) over neoadjuvant chemotherapy (NCT) in resectable esophageal carcinoma remains inconclusive. This study (NewEC) used global data to comprehensively evaluate these comparisons and to provide a preferable strategy for patient subsets. METHODS: This study included a meta-analysis of randomized controlled trials (RCTs) identified from inception to May 2019 from PubMed, EMBASE, the Cochrane Central Register of Controlled Trials, ClinicalTrials.gov, and congresses and a registry-based cohort study with patients from Massachusetts General Hospital (Massachusetts, USA) and Guangdong Provincial People's Hospital (Guangzhou, China) recruited from November 2000 and June 2017, to cross-validate the comparisons among NCRT versus NCT versus surgery. The GRADE approach was used to assessed quality of evidence in meta-analysis. Neural network machine learning propensity score-matched analysis was used to account for confounding by patient-level characteristics in the cohort study. The primary endpoint was overall survival (OS). The study was registered with PROSPERO CRD42017072242 and ClinicalTrials.gov NCT04027543. FINDINGS: Of 22,070 studies assessed, there were 38 (n = 6,993 patients) eligible RCTs. Additionally, 423 out of 467 screened patients were included in the cohort study. The results from trials showed that NCT had a better OS than surgery alone (hazard ratio [HR] 0·88, 95% confidence interval [CI] 0·79-0·98; high quality) and was only favorable for adenocarcinoma (HR 0·83, 95% CI 0·72-0·96; moderate quality). High-quality evidence showed a significantly better OS for NCRT than surgery alone (HR 0·74, 95% CI 0·66-0·82) for both adenocarcinoma (HR 0·73, 95% CI 0·62-0·86) and squamous cell carcinoma (SCC) (HR 0·73, 95% CI 0·65-0·83). The OS benefit of NCRT over NCT was seen in the pairwise (HR 0·78, 95% CI 0·62-0·99; high quality) and network (HR 0·82, 95% CI 0·72-0·93; high quality) meta-analyses, with similar results before (HR 0·60, 95% CI 0·40-0·91) and after (HR 0·44, 95% CI 0·25-0·77) matching in the cohort study, leading to a significantly increased 5-year OS rate in both adenocarcinoma and SCC before and after matching. The increased benefits from NCT or NCRT were not associated with the risk of 30-day or in-hospital mortality. INTERPRETATION: NewEC Study provided high-quality evidence supporting the survival benefits of NCRT or NCT over surgery alone, with NCRT presenting the greatest benefit for resectable esophageal carcinoma. FUNDING: National Science and Technology Major Project, the National Natural Science Foundation of China, the Natural Science Foundation of Guangdong Province, the Guangzhou Science and Technology Major Program, the Medical artificial intelligence project of Sun Yat-Sen Memorial Hospital, the Guangdong Science and Technology Department, the Guangdong Province Medical Scientific Research Foundation, and Guangdong Provincial People's Hospital Intermural Program.

N,4-Dimethyl-N-(4-nitro-benz-yl)benzene-sulfonamide.

In the title compound, C(15)H(16)N(2)O(4)S, there is a dihedral angle of 63.30 (8)° between the nitro-benzyl and benzene rings, which are separated by a sulfonamide unit The crystal packing is stabilized by a C-H⋯O inter-action.

Revisiting the structural basis and energetic landscape of susceptibility difference between HLA isotypes to allergic rhinitis.

The human leukocyte antigen class II (HLA II) molecules are implicated in the immunopathogenesis of allergic rhinitis (AR). The HLA II contains three allelic isotypes HLA-DR, -DQ, and -QP that exhibit considerably different susceptibility to AR. Here, we investigated the structural basis and energetic landscape of the susceptibility difference between the three HLA II isotypes to AR by combining computational analysis and experimental assay. Multiple sequence alignment revealed a low conservation among the three subtypes with sequence identity of ∼10% between them, suggesting that the peptide repertoires presented by HLA-DR, -DP and -DQ are not overlapped to each other, and they may be involved in different immune functions and dysfunctions. Structural analysis imparted that the antigenic peptides are rooted on the peptide-binding groove of HLA molecules and hold in a PPII-like helical conformation. Subsequently, the interaction behavior of 17 AR allergen-derived peptides with HLA-DR, -DP and -DQ was investigated using a statistics-based quantitative structure-activity relationship (QSAR) predictor. It was found a significant difference between the binding capabilities of these antigenic peptides to HLA-DR and to HLA-DP/-DQ; the former showed a generally higher affinity than the latter with p-value of 0.02 obtained from 2-tailed Student's t-test. The computational findings were then confirmed by HLA II-peptide stability assay, which demonstrated that the AR allergen-derived peptides have a high in vitro selectivity for HLA-DR over HLA-DP/-DQ. Thus, the HLA-DR isotype, rather than HLA-DP and -DQ, is expected to associate with the pathological process of AR.

Effect of acupuncture on rats with acute gouty arthritis inflammation: a metabonomic method for profiling of both urine and plasma metabolic perturbation.

Acute gouty arthritis is a common inflammation model with multiple pathogenic mechanisms seen in clinical practice, for which acupuncture may potentially be an alternative therapy. To investigate the effect of acupuncture on acute gouty arthritis and search for its mechanism, a metabonomic method was developed in this investigation. Acute gouty arthritis model rats were induced by monosodium urate (MSU) crystals. The urine and plasma samples were collected at several time points and the endogenous metabolites were analyzed by an ultra-performance liquid chromatography coupled with a mass spectrometry (UPLC-MS). Data were analyzed using principal components analysis (PCA) and partial least squares (PLS) analysis to compare metabolic profiles of MSU crystal-induced acute gouty arthritis rats with MSU crystal-induced acute gouty arthritis, treated with acupuncture rats. The results showed that acupuncture could restore the metabolite network that disturbed by MSU administration. Our study indicates that UPLC-MS-based metabonomics can be used as a potential tool for the investigation of biological effect of acupuncture on acute gouty arthritis.

Expression of neural receptors in mouse meibomian gland.

PURPOSE: To investigate gene and protein expression profiles of neural receptors found in the mouse meibomian gland. RNA and protein levels were determined for neuropeptide Y (NPY) receptor, vasoactive intestinal peptide (VIP) receptor, substance P (SP) receptor, and muscarinic cholinergic receptor (mAChR) subtypes M1-M5 in the mouse meibomian gland. METHODS: Frozen sections of Balb/c mouse eyelids were subjected to laser capture microdissection to isolate pure samples of meibomian gland ductal and acinar cells. Real-time polymerase chain reaction, immunolabeling, and Western blot analysis for SP receptor, VIP receptor, NPY receptor, and mAChR subtypes M1-M5 were performed on meibomian gland ductal and acinar cells. RESULTS: Expression of NPY1 receptor, VIP receptor 1, SP receptor, and all 5 mAChR subtypes was found in all meibomian gland ductal and acinar cells analyzed by real-time polymerase chain reaction. Immunolabeling and Western blot analysis confirmed the presence of NPY1 receptor, VIP receptor 1, SP receptor, and all 5 mAChR subtypes in the meibomian gland. The levels were variable with the duct showing greater levels of NPY1 receptor, SP receptor, and mAChRs 1, 2, 4, and 5 than with the gland. CONCLUSIONS: VIP receptor 1, SP receptor, NPY1 receptor, and mAChR subtypes may be involved in the regulation of meibomian gland secretion. Laser capture microdissection in conjunction with gene expression analysis provides an excellent approach for studying meibomian gland cells about which relatively little is known at the molecular level.