James A. Stefater

Poor validity of residual volumes as a marker for risk of aspiration in critically ill patients.

Background and Aims:Elevated residual volumes (RV), considered a marker for the risk of aspiration, are used to regulate the delivery of enteral tube feeding. We designed this prospective study to validate such use. Methods:Critically ill patients undergoing mechanical ventilation in the medical, coronary, or surgical intensive care units in a university-based tertiary care hospital, placed on intragastric enteral tube feeding through nasogastric or percutaneous endoscopic gastrostomy tubes, were included in this study. Patients were fed Probalance (Nestlé USA) to provide 25 kcal/kg per day (to which 109 yellow microscopic beads and 4.5 mL of blue food coloring per 1,500 mL was added). Patients were randomized to one of two groups based on management of RV: cessation of enteral tube feeding for RV >400 mL in study patients or for RV >200 mL in controls. Acute Physiology and Chronic Health Evaluation (APACHE) III, bowel function score, and aspiration risk score were determined. Bedside evaluations were done every 4 hrs for 3 days to measure RV, to detect blue food coloring, to check patient position, and to collect secretions from the trachea and oropharynx. Aspiration/regurgitation events were defined by the detection of yellow color in tracheal/oropharyngeal samples by fluorometry. Analysis was done by analysis of variance, Spearman’s correlation, Student’s t-test, Tukey’s method, and Cochran-Armitage test. Results:Forty patients (mean age, 44.6 yrs; range, 18–88 yrs; 70% male; mean APACHE III score, 40.9 [range, 12–85]) were evaluated (21 on nasogastric, 19 on percutaneous endoscopic gastrostomy feeds) and entered into the study. Based on 1,118 samples (531 oral, 587 tracheal), the mean frequency of regurgitation per patient was 31.3% (range, 0% to 94%), with a mean RV for all regurgitation events of 35.1 mL (range, 0–700 mL). The mean frequency of aspiration per patient was 22.1% (range, 0% to 94%), with a mean RV for all aspiration events of 30.6 mL (range, 0–700 mL). The median RV for both regurgitation and aspiration events was 5 mL. Over a wide range of RV, increasing from 0 mL to >400 mL, the frequency of regurgitation and aspiration did not change appreciably. Aspiration risk and bowel function scores did not correlate with the incidence of aspiration or regurgitation. Blue food coloring was detected on only three of the 1,118 (0.27%) samples. RV was ≤50 mL on 84.1% and >400 mL on 1.4% of bedside evaluations. Sensitivities for detecting aspiration per designated RV were as follows: 400 mL = 1.5%; 300 mL = 2.3%; 200 mL = 3.0%; and 150 mL = 4.5%. Low RV did not assure the absence of events, because the frequency of aspiration was 23.0% when RV was <150 mL. Raising the designated RV for cessation of enteral tube feeding from 200 mL to 400 mL did not increase the risk, because the frequency of aspiration was no different between controls (21.6%) and study patients (22.6%). The frequency of regurgitation was significantly less for patients with percutaneous endoscopic gastrostomy tubes compared with those with nasogastric tubes (20.3% vs. 40.7%, respectively; p = .046). There was no correlation between the incidence of pneumonia and the frequency of regurgitation or aspiration. Conclusions:Blue food coloring should not be used as a clinical monitor. Converting nasogastric tubes to percutaneous endoscopic gastrostomy tubes may be a successful strategy to reduce the risk of aspiration. No appropriate designated RV level to identify aspiration could be derived as a result of poor sensitivity over a wide range of RV. Study results do not support the conventional use of RV as a marker for the risk of aspiration.

Regulation of angiogenesis by a non-canonical Wnt-Flt1 pathway in myeloid cells

Myeloid cells are a feature of most tissues. Here we show that during development, retinal myeloid cells (RMCs) produce Wnt ligands to regulate blood vessel branching. In the mouse retina, where angiogenesis occurs postnatally, somatic deletion in RMCs of the Wnt ligand transporter Wntless results in increased angiogenesis in the deeper layers. We also show that mutation of Wnt5a and Wnt11 results in increased angiogenesis and that these ligands elicit RMC responses via a non-canonical Wnt pathway. Using cultured myeloid-like cells and RMC somatic deletion of Flt1, we show that an effector of Wnt-dependent suppression of angiogenesis by RMCs is Flt1, a naturally occurring inhibitor of vascular endothelial growth factor (VEGF). These findings indicate that resident myeloid cells can use a non-canonical, Wnt–Flt1 pathway to suppress angiogenic branching.

Metchnikoff's policemen: macrophages in development, homeostasis and regeneration

Over the past decade, modern genetic tools have permitted scientists to study the function of myeloid lineage cells, including macrophages, as never before. Macrophages were first detected more than a century ago as cells that ingested bacteria and other microbes, but it is now known that their functional roles are far more numerous. In this review, we focus on the prevailing functions of macrophages beyond their role in innate immunity. We highlight examples of macrophages acting as regulators of development, tissue homoeostasis, remodeling (the reorganization or renovation of existing tissues) and repair. We also detail how modern genetic tools have facilitated new insights into these mysterious cells.

Myeloid WNT7b Mediates the Angiogenic Switch and Metastasis in Breast Cancer

Oncogenic targets acting in both tumor cells and tumor stromal cells may offer special therapeutic appeal. Interrogation of the Oncomine database revealed that 52 of 53 human breast carcinomas showed substantial upregulation of WNT family ligand WNT7B. Immunolabeling of human mammary carcinoma showed that WNT7B immunoreactivity was associated with both tumor cells and with tumor-associated macrophages. In the MMTV-PymT mouse model of mammary carcinoma, we found tumor progression relied upon WNT7B produced by myeloid cells in the microenvironment. Wnt7b deletion in myeloid cells reduced the mass and volume of tumors due to a failure in the angiogenic switch. In the tumor overall, there was no change in expression of Wnt/β-catenin pathway target genes, but in vascular endothelial cells (VEC), expression of these genes was reduced, suggesting that VECs respond to Wnt/β-catenin signaling. Mechanistic investigations revealed that failure of the angiogenic switch could be attributed to reduced Vegfa mRNA and protein expression in VECs, a source of VEGFA mRNA in the tumor that was limiting in the absence of myeloid WNT7B. We also noted a dramatic reduction in lung metastasis associated with decreased macrophage-mediated tumor cell invasion. Together, these results illustrated the critical role of myeloid WNT7B in tumor progression, acting at the levels of angiogenesis, invasion, and metastasis. We suggest that therapeutic suppression of WNT7B signaling might be advantageous due to targeting multiple aspects of tumor progression.

Macrophage Wnt-Calcineurin-Flt1 signaling regulates mouse wound angiogenesis and repair.

The treatment of festering wounds is one of the most important aspects of medical care. Macrophages are important components of wound repair, both in fending off infection and in coordinating tissue repair. Here we show that macrophages use a Wnt-Calcineurin-Flt1 signaling pathway to suppress wound vasculature and delay repair. Conditional mutants deficient in both Wntless/GPR177, the secretory transporter of Wnt ligands, and CNB1, the essential component of the nuclear factor of activated T cells dephosporylation complex, displayed enhanced angiogenesis and accelerated repair. Furthermore, in myeloid-like cells, we show that noncanonical Wnt activates Flt1, a naturally occurring inhibitor of vascular endothelial growth factor-A-mediated angiogenesis, but only when calcineurin function is intact. Then, as expected, conditional deletion of Flt1 in macrophages resulted in enhanced wound angiogenesis and repair. These results are consistent with the published link between enhanced angiogenesis and enhanced repair, and establish novel therapeutic approaches for treatment of wounds.

The Eyes Absent Proteins in Developmental and Pathological Angiogenesis

Management of neoangiogenesis remains a high-value therapeutic goal. A recently uncovered association between the DNA damage repair pathway and pathological angiogenesis could open previously unexplored possibilities for intervention. An attractive and novel target is the Eyes absent (EYA) tyrosine phosphatase, which plays a critical role in the repair versus apoptosis decision after DNA damage. This study examines the role of EYA in the postnatal development of the retinal vasculature and under conditions of ischemia-reperfusion encountered in proliferative retinopathies. We find that the ability of the EYA proteins to promote endothelial cell (EC) migration contributes to a delay in postnatal development of the retinal vasculature when Eya3 is deleted specifically in ECs. By using genetic and chemical biology tools, we show that EYA contributes to pathological angiogenesis in a model of oxygen-induced retinopathy. Both in vivo and in vitro, loss of EYA tyrosine phosphatase activity leads to defective assembly of γ-H2AX foci and thus to DNA damage repair in ECs under oxidative stress. These data reveal the potential utility of EYA tyrosine phosphatase inhibitors as therapeutic agents in inhibiting pathological neovascularization with a range of clinical applications.

Pink hypopyon in a patient with Serratia marcescens corneal ulceration

A 65-year-old woman presented to the emergency ward at the Massachusetts Eye and Ear Infirmary with 2 days of redness, irritation, photophobia, and diminished vision in her left eye. She was found to have a large central corneal ulcer with a small hypopyon. On the following day, after initiation of broad-spectrum antibiotics, the patient had improved symptoms but now had a 2-mm hypopyon that was distinctly pink in color. Cultures were positive for Serratia marcescens. A pink hypopyon, a rare occurrence, alerted the authors to a causative agent of Enterobacteriacae, either Klebsiella or Serratia. Immediate and intensive treatment was subsequently initiated.

Case 4: Zone I Pediatric Open Globe Management

A 4-year-old boy presented with a Zone I open globe injury of the left eye. The child was playing with other children at a campground when he was hit directly in the left eye by a wooden stick. The patient was seen at another hospital and transferred to Massachusetts Eye & Ear (MEE) due to concern for open-globe injury. On presentation, the patient was found to have a large, vertical, corneal laceration extending from 12 to 6:30 o’clock with iris prolapse and concern for possible lens violation. The patient was taken to the operating room for exploration and repair of his corneal laceration. During the post-operative period, vision gradually improved, and the lens capsule was noted to be intact. A small posterior subcapsular cataract was noted in the initial post-operative period. Sutures were removed intraoperatively after 1 month due to faster wound healing in most children compared to adults, and the vision continued to improve. The patient followed up with a pediatric and corneal specialist closer to home and continued standard amblyopia therapy with good results.

Case 31: Delayed Presentation Zone I/II Open Globe Injury

A 28-year-old man presented with a Zone I and II open globe injury of the left eye. Following workplace trauma, the patient presented to an outside urgent care facility and was diagnosed with a corneal abrasion. Four days later, continued worsening of his blurry vision prompted an outside hospital emergency room visit. The patient was ultimately transferred to MEE and underwent surgical repair for his delayed presentation open globe injury of his left eye. This case highlights the importance of maintaining a high level of suspicion of an open globe injury in the setting of trauma. Self-sealing corneal wounds can present in a delayed fashion and may be missed if obvious findings such as subconjunctival hemorrhage or uveal prolapse are not present.

Case 2: Linear Corneal Laceration from Broken Glass

A 24-year-old woman presented with a Zone I open globe injury of the left eye from a shattered glass door. During surgical repair of the open globe injury, retained small foreign bodies were identified and removed intraoperatively. The pre-operative and post-operative course was uncomplicated, resulting in excellent visual acuity. This case demonstrates the potential for excellent visual outcomes in cases of Zone I injury, as well as the importance of careful intraoperative exploration for potentially retained foreign body material.