Optimizing neurotransmitter pathway detection by IR-MALDESI-MSI in mouse brain.

Mass spectrometry imaging (MSI) platforms such as infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI) are advantageous for a variety of applications, including elucidating the localization of neurotransmitters (NTs) and related molecules with respect to ion abundance across a sample without the need for derivatization or organic matrix application. While IR-MALDESI-MSI conventionally uses a thin exogenous ice matrix to improve signal abundance, it has been previously determined that sucrose embedding without the ice matrix improves detection of lipid species in striatal, coronal mouse brain sections. This work considers components of this workflow to determine the optimal sample preparation and matrix to enhance the detection of NTs and their related metabolites in coronal sections from the striatal region of the mouse brain. The discoveries herein will enable more comprehensive follow-on studies for the investigation of NTs to enrich biological pathways and interpretation related to neurodegenerative diseases and ischemic stroke.

A tale of two tumors: differential, but detrimental, effects of glioblastoma extracellular vesicles (EVs) on normal human brain cells.

Glioblastomas (GBMs) are dreadful brain tumors with abysmal survival outcomes. GBM EVs dramatically affect normal brain cells (largely astrocytes) constituting the tumor microenvironment (TME). EVs from different patient-derived GBM spheroids induced differential transcriptomic, secretomic, and proteomic effects on cultured astrocytes/brain tissue slices as GBM EV recipients. The net outcome of brain cell differential changes nonetheless converges on increased tumorigenicity. GBM spheroids and brain slices were derived from neurosurgical patient tissues following informed consent. Astrocytes were commercially obtained. EVs were isolated from conditioned culture media by ultrafiltration, ultraconcentration, and ultracentrifugation. EVs were characterized by nanoparticle tracking analysis, electron microscopy, biochemical markers, and proteomics. Astrocytes/brain tissues were treated with GBM EVs before downstream analyses. EVs from different GBMs induced brain cells to alter secretomes with pro-inflammatory or TME-modifying (proteolytic) effects. Astrocyte responses ranged from anti-viral gene/protein expression and cytokine release to altered extracellular signal-regulated protein kinase (ERK1/2) signaling pathways, and conditioned media from EV-treated cells increased GBM cell proliferation. Thus, astrocytes/brain slices treated with different GBM EVs underwent non-identical changes in various 'omics readouts and other assays, indicating "personalized" tumor-specific GBM EV effects on the TME. This raises concern regarding reliance on "model" systems as a sole basis for translational direction. Nonetheless, net downstream impacts from differential cellular and TME effects still led to increased tumorigenic capacities for the different GBMs.

Digital assessment of supracrestal tissue attachment and its correlation with dentogingival components.

BACKGROUND: The aim of this study is to measure, in vivo, the supracrestal tissue attachment dimensions (STADs) by means of a noninvasive digital method and to investigate the association between STADs and gingival thickness (GT), tooth position, tooth length, tooth width, keratinized tissue width (KTW), buccal bone thickness (BBT), and bone crest (BC) level. METHODS: Nineteen periodontally healthy subjects who previously received full mouth periodontal charting, cone beam computed tomography, and intraoral scan for the purpose of implant planning were included in the study. A digital imaging software was used for the superimposition of Digital Imaging and Communications in Medicine and stereolithography files, along with hard and soft tissue measurements. Pearson's correlation and ANOVA statistical analyses were used to investigate potential trends between STADs and other dentogingival components. RESULTS: A total of 203 teeth were assessed, with an average STADs of 2.05 mm (±0.99 mm). STADs were larger in mandibular than maxillary teeth (p-value <0.001) and decreased from anterior to posterior teeth. STADs exhibited an inverse relationship with BBTs and GTs (p-value <0.001) and the KTW (p-value = 0.05). Positive correlations were found between GT and BBT (p-value <0.001), whereas both were negatively correlated with the distance between the cementoenamel junction and BC (p-values 0.019 and 0.006, respectively) and positively correlated with KTW (p-value <0.001). CONCLUSIONS: This study highlighted the dynamic nature of STA relative to tooth position. Additionally, it explored the intricate relationships of STADs with various dentogingival components. KEY POINTS: To the best of the authors' knowledge, this study represents the first application of CBCTs, intraoral scans, and clinical probe depths for noninvasive supracrestal tissue attachment measurements. This study advocates for a personalized assessment of supracrestal attachments, incorporating tooth position and other dentogingival components. The study emphasizes the importance for practitioners to consider the specific patient gingival phenotypes during restorative or surgical planning to avoid adverse outcomes.

Assessing Larval Zebrafish Survival and Gene Expression Following Sodium Butyrate Exposure and Subsequent Lethal Bacterial Lipopolysaccharide (LPS) Endotoxin Challenge.

As aquaculture production continues to grow, producers are looking for more sustainable methods to promote growth and increase fish health and survival. Butyrate is a short-chain fatty acid (SCFA) with considerable benefits to gut health, and in recent years, butyrate has been commonly used as an alternative to antimicrobials in livestock production. In this study, we aimed to assess the protective effects of sodium butyrate (NaB) on larval zebrafish subjected to a lethal Pseudomonas aeruginosa lipopolysaccharide (LPS) endotoxin challenge and to elucidate potential protective mechanisms of action. Larval zebrafish were pre-treated with 0, 3000, or 6000 µM NaB for 24 h at 72 h post-fertilization (hpf), then immune challenged for 24 h with 60 µg/mL of LPS at 96 hpf. Our results demonstrate that larval zebrafish pre-treated with 6000 µM of NaB prior to lethal LPS challenge experienced significantly increased survival by 40%, and this same level of NaB significantly down-regulated the expression of pro-inflammatory Tumor Necrosis Factor α (TNF-alpha). Findings from this study are consistent with the beneficial effects of NaB on other vertebrate species and support the potential use of NaB in aquaculture.

Comparative analysis of sucrose-embedding for whole-body zebrafish MSI by IR-MALDESI.

Infrared matrix-assisted laser desorption electrospray ionization mass spectrometry imaging (IR-MALDESI) conventionally utilizes fresh-frozen biological tissues with an ice matrix to improve the detection of analytes. Sucrose-embedding with paraformaldehyde fixation has demonstrated feasibility as an alternative matrix for analysis by IR-MALDESI by preserving tissue features and enhancing ionization of lipids. However, investigating multi-organ systems provides broader context for a biological study and can elucidate more information about a disease state as opposed to a single organ. Danio rerio, or zebrafish, are model organisms for various disease states and can be imaged as a multi-organ sample to analyze morphological and metabolomic preservation as a result of sample preparation. Herein, whole-body zebrafish were imaged to compare sucrose-embedding with paraformaldehyde fixation against conventional fresh-frozen sample preparation. Serial sections were analyzed with and without an ice matrix to evaluate if sucrose functions as an alternative energy-absorbing matrix for IR-MALDESI applications across whole-body tissues. The resulting four conditions were compared in terms of total putative lipid annotations and category diversity, coverage across the entire m/z range, and ion abundance. Ultimately, sucrose-embedded zebrafish had an increase in putative lipid annotations for the combination of putative annotations with and without the application of an ice matrix relative to fresh-frozen tissues which require the application of an ice matrix. Upon the use of an ice matrix, a greater number of high mass putative lipid annotations (e.g., glycerophospholipids, glycerolipids, and sphingolipids) were identified. Conversely, without an ice matrix, sucrose-embedded sections elucidated more putative annotations in lower molecular weight lipids, including fatty acyls and sterol lipids. Similar to the mouse brain model, sucrose-embedding increased putative lipid annotation and abundance for whole-body zebrafish.

Prophylactic Tranexamic Acid Prevents Postpartum Hemorrhage and Transfusions in Cesarean Deliveries: A Systematic Review and Meta-analysis.

Postpartum hemorrhage (PPH) is the leading cause of maternal mortality worldwide and PPH resulting in transfusion is the most common maternal morbidity in the United States. Literature demonstrates that tranexamic acid (TXA) can reduce blood loss in cesarean deliveries; however, there is little consensus on the impact on major morbidities like PPH and transfusions. We conducted a systematic review/meta-analysis of randomized controlled trials (RCTs) to evaluate if administration of prophylactic intravenous (IV) TXA prevents PPH and/or transfusions following low-risk cesarean delivery. PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-analyses) guidelines were followed. Five databases were searched: Cochrane, EBSCO, Ovid, PubMed, and ClinicalKey. RCTs published in English between January 2000 and December 2021 were included. Studies compared PPH and transfusions in cesarean deliveries between prophylactic IV TXA and control (placebo or no placebo). The primary outcome was PPH, and the secondary outcome was transfusions. Random effects models were used to calculate effect size (ES) of exposure in Mantel-Haenszel risk ratios (RR). All analysis was done at a confidence level (CI) of α = 0.5. Modeling showed that TXA led to significantly less risk of PPH than control (RR: 0.43; 95% CI: 0.28-0.67). The effect on transfusion was comparable (RR: 0.39; 95% CI: 0.21-0.73). Heterogeneity was minimal (I 2 = 0%). Due to the large sample sizes needed, many RCTs are not powered to interpret TXA's effect on PPH and transfusions. Pooling these studies in a meta-analysis allows for more power and analysis but is limited by the heterogeneity of studies. Our results minimize heterogeneity while demonstrating that prophylactic TXA can lower PPH occurrence and reduce the need for blood transfusion. We suggest considering prophylactic IV TXA as the standard of care in low-risk cesarean deliveries. KEY POINTS: · Consider TXA prior to incision for singleton, term pregnancies undergoing elective cesarean.. · Prophylactic TXA is effective in preventing PPH and blood transfusions.. · Routine use of TXA has the potential to decrease transfusion-related complications and costs..

Time of acquisition and high spatial resolution mass spectrometry imaging.

The field of mass spectrometry imaging (MSI) is constantly evolving to analyze a diverse array of biological systems. A common goal is the need to resolve cellular and subcellular heterogeneity with high spatial resolution. As the field continues to progress towards high spatial resolution, other parameters must be considered when developing a practical method. Here, we discuss the impacts of high spatial resolution on the time of acquisition and the associated implications they have on an MSI analysis (e.g., area of the region of interest). This work presents a brief tutorial serving to evaluate high spatial resolution MSI relative to time of acquisition and data file size.

Lipidomic Analysis of Mouse Brain to Evaluate the Efficacy and Preservation of Different Tissue Preparatory Techniques by IR-MALDESI-MSI.

Numerous preparatory methods have been developed to preserve the cellular and structural integrity of various biological tissues for different -omics studies. Herein, two preparatory methods for mass spectrometry imaging (MSI) were evaluated, fresh-frozen and sucrose-embedded, paraformaldehyde (PFA) fixed, in terms of ion abundance, putative lipid identifications, and preservation of analyte spatial distributions. Infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI)-MSI was utilized to compare the preparatory methods of interest with and without the use of the conventional ice matrix. There were 2.5-fold and 1.6-fold more lipid species putatively identified in positive- and negative-ion modes, respectively, for sucrose-embedded, PFA-fixed tissues without an ice matrix relative to the current IR-MALDESI-MSI gold-standard, fresh-frozen tissue preparation with an exogenous ice matrix. Furthermore, sucrose-embedded tissues demonstrated improved spatial distribution of ions resulting from the cryo-protective property of sucrose and paraformaldehyde fixation. Evidence from these investigations supports sucrose-embedding without ice matrix as an alternative preparatory technique for IR-MALDESI-MSI.

Synthesis of Nε-acetyl-L-homolysine by the Lossen rearrangement and its application for probing deacetylases and binding modules of acetyl-lysine.

Lysine acetylation is a posttranslational protein modification mediating protein-protein interactions by recruitment of bromodomains. Investigations of bromodomains have focused so far on the sequence context of the modification site and acyl-modifications installed at lysine side chains. In contrast, there is only little information about the impact of the lysine residue that carries the modification on bromodomain binding. Here, we report a synthesis strategy for L-acetyl-homolysine from L-2-aminosuberic acid by the Lossen rearrangement. Peptide probes containing acetylated homolysine, lysine, and ornithine were generated and used for probing the binding preferences of four bromodomains from three different families. Tested bromodomains showed distinct binding patterns, and one of them bound acetylated homolysine with similar efficiency as the native substrate containing acetyl-lysine. Deacetylation assays with a bacterial sirtuin showed a strong preference for acetylated lysine, despite a broad specificity for N-acyl modifications.

Impact of osteopathic manipulative techniques on the management of dizziness caused by neuro-otologic disorders: systematic review and meta-analysis.

CONTEXT: Osteopathic manipulative treatment (OMT) has been utilized by osteopathic clinicians as primary or adjunctive management for dizziness caused by neuro-otologic disorders. To our knowledge, no current systematic reviews provide pooled estimates that evaluate the impact of OMT on dizziness. OBJECTIVES: We aimed to systematically evaluate the effectiveness and safety of OMT and analogous techniques in the treatment of dizziness. METHODS: We performed a literature search in CINAHL, Embase, MEDLINE, Allied and Complementary Medicine Database (AMED), EMCare, Physiotherapy Evidence Database (PEDro), PubMed, PsycINFO, Osteopathic Medicine Digital Library (OSTMED.DR), and Cochrane Central Register of Controlled Trials (CENTRAL) from inception to March 2021 for randomized controlled trials (RCTs) and prospective or retrospective observational studies of adult patients experiencing dizziness from neuro-otological disorders. Eligible studies compared the effectiveness of OMT or OMT analogous techniques with a comparator intervention, such as a sham manipulation, a different manual technique, standard of care, or a nonpharmacological intervention like exercise or behavioral therapy. Assessed outcomes included disability associated with dizziness, dizziness severity, dizziness frequency, risk of fall, improvement in quality of life (QOL), and return to work (RTW). Assessed harm outcomes included all-cause dropout (ACD) rates, dropouts due to inefficacy, and adverse events. The meta-analysis was based on the similarities between the OMT or OMT analogous technique and the comparator interventions. The risk of bias (ROB) was assessed utilizing a modified version of the Cochrane Risk of Bias Tool for RCTs and the Cochrane Risk of Bias in Non-randomized Studies - of Interventions (ROBINS-I) for observational studies. The quality of evidence was determined utilizing the Grading of Recommendations, Assessment, Development, and Evaluations (GRADE) approach. RESULTS: There were 3,375 studies identified and screened, and the full text of 47 of them were reviewed. Among those, 12 (11 RCTs, 1 observational study, n=367 participants) met the inclusion criteria for data extraction. Moderate-quality evidence showed that articular OMT techniques were associated with decreases (all p<0.01) in disability associated with dizziness (n=141, mean difference [MD]=-11, 95% confidence interval [CI]=-16.2 to -5.9), dizziness severity (n=158, MD=-1.6, 95% CI=-2.4 to -0.7), and dizziness frequency (n=136, MD=-0.6, 95% CI=-1.1 to -0.2). Low-quality evidence showed that articular OMT was not associated with ACD rates (odds ratio [OR]=2.2, 95% CI=0.5 to 10.2, p=0.31). When data were pooled for any type of OMT technique, findings were similar; however, disability associated with dizziness and ACD rates had high heterogeneity (I2=59 and 46%). No studies met all of the criteria for ROB. CONCLUSIONS: The current review found moderate-quality evidence that treatment with articular OMT techniques was significantly associated with decreased disability associated with dizziness, dizziness severity, and dizziness frequency. However, our findings should be interpreted cautiously because of the high ROB and small sample sizes in the eligible studies.

What Happened and Why: Responding to Racism, Discrimination, and Microaggressions in the Clinical Learning Environment.

Introduction: Within clinical learning environments, medical students are uniquely faced with power differentials that make acts of racism, discrimination, and microaggressions (RDM) challenging to address. Experiences of microaggressions and mistreatment are correlated with higher rates of positive depression screening and lower satisfaction with medical training. We developed a curriculum for medical students beginning clerkship rotations to promote the recognition of and response to RDM. Methods: Guided by generalized and targeted needs assessments, we created a case-based curriculum to practice communication responses to address RDM. The communication framework, a 6Ds approach, was developed through adaptation and expansion of established and previously learned communication upstander frameworks. Cases were collected through volunteer submission and revised to maintain anonymity. Faculty and senior medical students cofacilitated the small-group sessions. During the sessions, students reviewed the communication framework, explored their natural response strategies, and practiced all response strategies. Results: Of 196 workshop participants, 152 (78%) completed the evaluation surveys. Pre- and postsession survey cohort comparison demonstrated a significant increase in students' awareness of instances of RDM (from 34% to 46%), knowledge of communication strategies to mitigate RDM (presession M = 3.4, postsession M = 4.6, p < .01), and confidence to address RDM (presession M = 3.0, postsession M = 4.4, p < .01). Discussion: Students gained valuable communication skills from interactive sessions addressing RDM using empathy, reflection, and relatability. The workshop empowered students to feel prepared to enter professional teams and effectively mitigate harmful discourse.

Epithelial-interleukin-1 inhibits collagen formation by airway fibroblasts: Implications for asthma.

In asthma, the airway epithelium has an impaired capacity to differentiate and plays a key role in the development of airway inflammation and remodeling through mediator release. The study objective was to investigate the release of (IL)-1 family members from primary airway epithelial-cells during differentiation, and how they affect primary airway fibroblast (PAF)-induced inflammation, extracellular matrix (ECM) production, and collagen I remodeling. The release of IL-1α/ß and IL-33 during airway epithelial differentiation was assessed over 20-days using air-liquid interface cultures. The effect of IL-1 family cytokines on airway fibroblasts grown on collagen-coated well-plates and 3-dimensional collagen gels was assessed by measurement of inflammatory mediators and ECM proteins by ELISA and western blot, as well as collagen fiber formation using non-linear optical microscopy after 24-hours. The production of IL-1α is elevated in undifferentiated asthmatic-PAECs compared to controls. IL-1α/ß induced fibroblast pro-inflammatory responses (CXCL8/IL-8, IL-6, TSLP, GM-CSF) and suppressed ECM-production (collagen, fibronectin, periostin) and the cell's ability to repair and remodel fibrillar collagen I via LOX, LOXL1 and LOXL2 activity, as confirmed by inhibition with ß-aminopropionitrile. These data support a role for epithelial-derived-IL-1 in the dysregulated repair of the asthmatic-EMTU and provides new insights into the contribution of airway fibroblasts in inflammation and airway remodeling in asthma.

Sensory focused exercise improves anxiety in Parkinson's disease: A randomized controlled trial.

Anxiety has been implicated as one of the greatest influences on quality of life in Parkinson's disease (PD). The etiology of anxiety is unclear, although previous work suggests that anxiety may be linked to sensory deficits that cause uncertainty in movement. Thus, the current study examined whether focusing attention on sensory feedback during goal-based exercise has the potential to provide benefits to anxiety in PD. Thirty-five participants with PD were randomized to either a Sensory Attention Focused Exercise (SAFEx) (i.e. internal focus of attention, n = 18) or Sham Exercise control (i.e. external focus of attention, n = 17) and completed 33 one-hour attention-based exercise sessions over 11-weeks. Before and after the program (pre and post), participants completed the Parkinson Anxiety Scale (PAS) questionnaire. The PAS includes three anxiety sections: persistent, episodic, and avoidance. Changes in the total PAS score and within each section of the PAS were subjected to two-factor mixed repeated measures ANCOVA. Significant group by time interactions demonstrated that from pre to post, total PAS scores (p = 0.007) and episodic anxiety scores (p = 0.010) significantly decreased in the SAFEx group only (ΔTotal PAS = -5.2, F(1,27) = 5.41, p = 0.028, ηp2 = 0.17; ΔEpisodic Score = -1.8, F(1,27) = 6.89, p = 0.014, ηp2 = 0.20). In conclusion, focusing attention on sensory feedback while completing goal-based exercises may provide significant benefits to improving anxiety in PD. As such, sensory attention focused exercise may be a critical adjunct therapy for improving anxiety, and ultimately quality of life in people with PD.

Frailty as a Predictor of Neurosurgical Outcomes in Brain Tumor Patients.

BACKGROUND: Preoperative risk assessment is important, but inexact because physiologic reserves are difficult to measure. When assessing quality of life for patients with brain tumors, having a better predictor of postsurgical outcome would be beneficial in counseling these patients. Frailty is thought to estimate physiologic reserves, and it has been found to predict postoperative complications, length of stay, and discharge to a skilled nursing facility or assisted living facility in patients undergoing various types of surgery. Frailty as an adjunct to preoperative assessment of neurosurgical patients has never been evaluated. This study aimed to determine whether frailty predicts neurosurgical complications in patients with brain tumors and enhances current perioperative risk models. METHODS: Frailty was preoperatively assessed in 260 patients undergoing surgery for brain tumor resection using a validated scale that assessed weakness, weight loss, exhaustion, low physical activity, and slowed walking speed. Patients were classified as nonfrail (score of 0-1), moderately frail (score of 2-3), or frail (score of 4-5). Moderately frail and frail patients were combined for analysis. RESULTS: Preoperative frailty was associated with an increased risk for discharge to a location other than home (10.36; 95% confidence interval, 3.6-30.1), postoperative complications (2.09; 95% confidence interval, 1.09-3.98), and a longer length of stay (1.66; 95% confidence interval, 1.24-2.21). CONCLUSIONS: Frailty independently predicts discharge disposition, postoperative complications, and length of stay in patients undergoing surgery for brain tumor resection. Preoperative assessment of frailty can help neurosurgeons and patients make more informed decisions about pursing surgical treatment.

The Incidence and Durability of Compensatory Hypertrophy in Pediatric Patients with Solitary Kidneys.

OBJECTIVE: To evaluate the incidence and durability of compensatory hypertrophy with solitary kidneys in the setting of those with multicystic dysplastic kidney (MCDK) or Wilms tumor (WT) status postnephrectomy. PATIENTS AND METHODS: We conducted a retrospective cohort study of patients with MCDK and WT. MCDK patients were verified by sonographic findings prenatally. WT patients entered our study at time of nephrectomy. We compared the natural history of hypertrophy between the 2 cohorts via renal length measurement. We performed linear regression to predict creatinine clearance from renal length after adjusting for age and cohort status (MCDK versus WT). RESULTS: Fifty-two patients (56%) were diagnosed with WT, and 71 patients (44%) with MCDK patients met study criteria with a median age of postnatal ultrasound at 1.6 months (interquartile range 0.5-3.6). The median (IQR) follow-up time was 7.5 years. At the time of nephrectomy, 25/52 (48%) of Wilms patients had contralateral hypertrophy, while at diagnosis 22/71 (31%) of MCDK patients had contralateral hypertrophy, P = .03. Contralateral hypertrophy was a consistent finding throughout follow-up. As renal length increases by 1 cm, glomerular filtration rate increased by 7.8 mL/min/m2 (95% confidence interval 1.8-13.8, P = .01). CONCLUSION: Contralateral hypertrophy appears to be a compensatory mechanism for unilateral kidney pathology as demonstrated by MCDK and WT patients. Hypertrophy often occurs at the time of diagnosis and appears to be a permanent finding as children reach their teenage years. Additionally, in nonhydronephrotic kidneys, increases in renal length correlate with improvement in glomerular filtration rate. Overall, the majority of children with solitary kidneys demonstrate compensatory hypertrophy.

Research and management of plastic pollution in coastal environments of China.

Marine plastic waste has become an ever-increasing environmental threat in the world's ocean largely due to their unique properties and ubiquitous occurrence. They include diverse forms of land- and ocean-based sources of plastics and are estimated to account for up to 85% of marine debris worldwide. As secondary pollutants, marine microplastic particles (<5 mm) are derived from pellet loss and degradation of macroplastics. Up to now, several reports have proposed negative impacts of both macro-sized and micro-sized plastics on marine biota. As one of the rapidly growing economies, China is the topmost contributor of plastic waste in the world. China's massive impact on the plastic levels of the ocean are a definite cause of concern and is developing multiple economic, environmental and biological complications. The research of plastics impact on coastal environments in China is only incipient. Here we review the available information on plastic waste, their impacts on marine biota and human health, and Chinese government policies and management initiatives. Although Chinese coastal environments (surface water, coastal sediments, water column) are affected by microplastics pollution, both from land-based and sea-based activities, their impacts on marine biota remain to be elucidated. Though national-level policies are modern and well suited for minimizing the impacts of plastic pollution, there is hardly any legislation for containment of microplastic pollution. Our objective is to review and summarize the information about the occurrence, impacts, and management of plastic pollution in the Chinese coastal environments in order to comprehend their widespread repercussions. MAIN FINDING: Microplastics are increasingly being detected and quantified in Chinese coastal environments and legislation for containment of such pollution is highly recommended.

Augmented Weighted Estimators Dealing with Practical Positivity Violation to Causal inferences in a Random Coefficient Model.

The inverse probability of treatment weighted (IPTW) estimator can be used to make causal inferences under two assumptions: (1) no unobserved confounders (ignorability) and (2) positive probability of treatment and of control at every level of the confounders (positivity), but is vulnerable to bias if by chance, the proportion of the sample assigned to treatment, or proportion of control, is zero at certain levels of the confounders. We propose to deal with this sampling zero problem, also known as practical violation of the positivity assumption, in a setting where the observed confounder is cluster identity, i.e., treatment assignment is ignorable within clusters. Specifically, based on a random coefficient model assumed for the potential outcome, we augment the IPTW estimating function with the estimated potential outcomes of treatment (or of control) for clusters that have no observation of treatment (or control). If the cluster-specific potential outcomes are estimated correctly, the augmented estimating function can be shown to converge in expectation to zero and therefore yield consistent causal estimates. The proposed method can be implemented in the existing software, and it performs well in simulated data as well as with real-world data from a teacher preparation evaluation study.

Mechanical stretch upregulates connexin43 in human trabecular meshwork cells.

BACKGROUND: Primary open angle glaucoma (POAG) patients have hallmark increases in intraocular pressure (IOP) and noted dysfunction of the trabecular meshwork (TM). Connexin43 (Cx43) is a gap junction widely expressed on the TM that is important for intercellular communication. The human gene is known as gap junction alpha-1 (GJA1). Since the role of Cx43 in the TM is not fully understood, we set out to determine the effect of excess mechanical stretch on cultured human trabecular meshwork cells (hTMCs) and to specifically investigate the effect of stretch on Cx43 expression and function. METHODS: Primary hTMCs were cultured and subjected to 48 hours of 15% cyclic mechanical stretch at a frequency of 1 Hz. Levels of apoptosis and necrosis secondary to stretch were investigated using colorimetric assays. The effect of stretch on gap junction Cx43 and GJA1 was investigated by RT-PCR, immunoblotting and immunofluorescence. The migration of Lucifer Yellow dye was used to assess intercellular communication. RESULTS: Stretch significantly increased the rates of apoptosis and necrosis in hTMCs. The increased rate of injury in stretched hTMCs was further associated with significant upregulation of GJA1 mRNA and Cx43 protein. Upregulation of Cx43 protein was concomitant to increased intercellular communication. CONCLUSIONS: We have shown stretch to increase GJA1 gene and Cx43 protein expression, as well as intercellular communication. We have further shown stretch to be injurious to hTMCs. Upregulation of Cx43 in the hTM subsequent to stretch is a novel finding, which may be useful in elucidating the mechanism of TM injury in POAG patients.

Benefit of Intracystic Bleomycin for Symptomatic Recurrent Rathke Cleft Cyst.

BACKGROUND: Management of recurrent Rathke cleft cysts (RCC) is problematic. The mainstay of treatment has been reoperation with cyst drainage. Radical cyst resection, although effective, results in a high incidence of diabetes insipidus. Several case reports suggest a potential benefit to radiation therapy or the use of intracystic bleomycin. The intracystic application of bleomycin is known to be beneficial in the treatment of cystic craniopharyngioma; however, its usefulness in the treatment of recurrent RCC has yet to be proven. OBJECTIVE: To present our 6-yr experience using intracystic bleomycin for recurrent RCC. METHODS: We performed a retrospective chart review of patients with RCC who underwent surgical resection between January 2010 and May 2016 by a single surgeon. Specific attention was paid to patients with recurrent RCC. RESULTS: Of the 59 patients operated on for RCC during this 6-yr interval, 6 patients with symptomatic recurrent RCC were identified and received intracystic bleomycin at the time of reoperation. To date, no symptomatic cyst recurrence has been documented in the patients receiving bleomycin (mean = 38.8 mo, range 21.2-79.8 mo). CONCLUSION: The use of intracystic bleomycin appears to be a safe and potentially effective treatment option in patients with recurrent RCC. Additional studies with longer follow-up are needed to further define the role of bleomycin in recurrent RCC.