Genetically engineered and enucleated human mesenchymal stromal cells for the targeted delivery of therapeutics to diseased tissue.

Targeting the delivery of therapeutics specifically to diseased tissue enhances their efficacy and decreases their side effects. Here we show that mesenchymal stromal cells with their nuclei removed by density-gradient centrifugation following the genetic modification of the cells for their display of chemoattractant receptors and endothelial-cell-binding molecules are effective vehicles for the targeted delivery of therapeutics. The enucleated cells neither proliferate nor permanently engraft in the host, yet retain the organelles for energy and protein production, undergo integrin-regulated adhesion to inflamed endothelial cells, and actively home to chemokine gradients established by diseased tissues. In mouse models of acute inflammation and of pancreatitis, systemically administered enucleated cells expressing two types of chemokine receptor and an endothelial adhesion molecule enhanced the delivery of an anti-inflammatory cytokine to diseased tissue (with respect to unmodified stromal cells and to exosomes derived from bone-marrow-derived stromal cells), attenuating inflammation and ameliorating disease pathology. Enucleated cells retain most of the cells' functionality, yet acquire the cargo-carrying characteristics of cell-free delivery systems, and hence represent a versatile delivery vehicle and therapeutic system.

Comparison of Serial Blood Collection by Facial Vein and Retrobulbar Methods in C57BL/6 Mice.

Many biomedical research protocols for mouse models involve serial blood collection and analysis. Two common techniques for serial blood collection in this species are the retrobulbar (RB, also called retroorbital) and facial vein (FV) methods. However, previous studies comparing these methods typically evaluated collection at a maximum of 2 time points. Here we compared hematologic values, adverse clinical effects, and histopathologic lesions in mice bled either once or serially (6 times) by using the FV or RB method. Mice (n = 48) were divided into 4 groups: single FV, single RB, serial FV and serial RB. Mice in the single-collection groups underwent a single blood collection by the indicated method, whereas those in the serial-collection groups were sampled once weekly for 6 consecutive weeks. All animals were euthanized and necropsied 2 wk after their last blood collection. Compared with all other groups, the serial FV group experienced more serious clinical adverse events, including 33% mortality, convulsions, head tilt, and hemorrhage from the ear canal and nares. In addition, mice in the FV groups had a significantly greater acute body weight loss compared with mice in the RB groups. Histologically, mice in both serial-collection groups had an increased incidence of tissue lesions compared with their respective single-collection groups. Importantly, only mice in the serial FV group had life-threatening histopathologic lesions, including cerebral hemorrhage or ischemia. Given these data, we conclude that serial blood collection in mice causes increased incidence of tissue damage compared with single sampling, and serial blood collection by the FV method causes substantial morbidity and mortality compared with the RB method.

Outcome and clinical, pathological, and immunohistochemical factors associated with prognosis for dogs with early-stage anal sac adenocarcinoma treated with surgery alone: 34 cases (2002-2013).

OBJECTIVE To determine survival time and metastatic rate for dogs with early-stage anal sac adenocarcinoma (ASACA) treated with surgery alone and assess whether specific clinical, pathological, or immunohistochemical factors were predictive of outcome for those dogs. DESIGN Retrospective case series. ANIMALS 34 dogs with early-stage, nonmetastatic ASACA that were treated with surgery only. PROCEDURES Medical record databases of 2 referral hospitals were searched to identify dogs examined between 2002 and 2013 that had a diagnosis of nonmetastatic ASACA that was < 3.2 cm at its largest diameter. Only dogs that received surgical treatment alone were included in the study. For each dog, information extracted from the medical record included signalment, clinical and diagnostic test findings, tumor characteristics, and outcome. When available, archived tumor specimens were histologically reviewed and tumor characteristics were described; Ki-67 and E-cadherin expressions were evaluated by use of immunohistochemical methods. Clinical, pathological, and immunohistochemical factors were assessed for associations with survival time and tumor recurrence and metastasis rates. RESULTS Median survival time was 1,237 days. Seven dogs had tumor recurrence and 9 dogs developed metastatic disease at a median of 354 and 589 days, respectively, after primary tumor removal. Cellular pleomorphism was positively associated with development of metastatic disease. No other factors evaluated were associated with outcome. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated dogs with early-stage nonmetastatic ASACA generally had a favorable outcome following surgical removal of the primary tumor alone. Routine rectal examination may be a simple and useful method for detection of dogs with early-stage ASACA.

Apical Elongation of Molar Teeth in Captive Microtus Voles.

Molar apical elongation (MAE) was the leading cause for euthanasia or death in a captive breeding colony of endangered Amargosa voles ( Microtus californicus scirpensis). Clinical signs included ocular discharge, abnormal mastication, dyspnea, abnormal mentation, weight loss, and death. Although the severity varied, all molars in all quadrants were affected. When severe, the overgrown molar reserve crown and apex protruded into the nasal meatuses, the orbit, the calvarial vault and through the ventral margin of the mandible. Overall prevalence in the colony was 63% (92/146 voles) and increased to 77% in aged voles (>1 year). Mean age of onset was 5.3 months (1.7-11.2 months). Progression to extreme severity occurred over 1 to 3 months. Mean survival was 10.9 months (7.1-21.7 months). Histologically, the lesion was characterized by odontogenic hyperplasia and dysplasia. MAE was also documented in museum specimens of 2 other M. californicus subspecies ( M. californicus californicus, M. californicus vallicola) and 3 other Microtus species ( M. montanus, M. pennsylvanicus, M. socialis). In the M. californicus californicus collection, overall prevalence was 35.1% (129/368 skulls) and increased to 77.3% in aged voles (>1 year). A probable genetic influence was identified in the museum collection of M. californicus californicus. The etiopathogenesis of MAE is likely multifactorial, due to (1) inherent continuous odontogenic proliferation, (2) inadequate occlusal attrition, and (3) possible heritable disease susceptibility. In captivity, dietary or other management of occlusal attrition to prevent or delay MAE is a fundamental concern.