Cross-Cutting Lessons Learned During the COVID-19 Pandemic-the Walter Reed Army Institute of Research Experience.

INTRODUCTION: At the start of the coronavirus disease 2019 (COVID-19) pandemic, Walter Reed Army Institute of Research (WRAIR) mobilized to rapidly conduct medical research to detect, prevent, and treat the disease in order to minimize the impact of the pandemic on the health and readiness of U.S. Forces. WRAIR's major efforts included the development of the Department of Defense (DoD) COVID-19 vaccine candidate, researching novel drug therapies and monoclonal antibodies, refining and scaling-up diagnostic capabilities, evaluating the impact of viral diversity, assessing the behavioral health of Soldiers, supporting U.S. DoD operational forces overseas, and providing myriad assistance to allied nations. WRAIR personnel have also filled key roles within the whole of government response to the pandemic. WRAIR had to overcome major pandemic-related operational challenges in order to quickly execute a multimillion-dollar portfolio of COVID-19 research. Consequently, the organization learned lessons that could benefit other leaders of medical research organizations preparing for the next pandemic. MATERIALS AND METHODS: We identified lessons learned using a qualitative thematic analysis of 76 observation/recommendation pairs from across the organization. These lessons learned were organized under the Army's four pillars of readiness (staffing, training, equipping, and leadership development). To this framework, we added organizing and leading to best capture our experiences within the context of pandemic response. RESULTS: The major lessons learned for organizing were: (1) the pandemic created a need to rapidly pivot to new scientific priorities; (2) necessary health and safety precautions disrupted the flow of normal science and put programs at risk of missing milestones; (3) relationships with partners and allies facilitated medical diplomacy and advancement of U.S. national military and economic goals; and (4) a successful response required interoperability within and across multiple organizations. For equipping: (1) existing infrastructure lacked sufficient capacity and technical capability to allow immediate countermeasure development; (2) critical supply chains were strained; and (3) critical information system function and capacity were suddenly insufficient under maximum remote work. For staffing and training: (1) successful telework required rapid shifts in management, engagement, and accountability methods; and (2) organizational policies and processes had to adapt quickly to support remote staffing. For leading and leadership development (1) engaged, hopeful, and empathetic leadership made a difference; and (2) the workforce benefitted from concerted leadership communication that created a shared understanding of shifting priorities as well as new processes and procedures. CONCLUSIONS: An effective pandemic response requires comprehensive institutional preparedness that facilitates flexibility and surge capacity. The single most important action leaders of medical research organizations can take to prepare for the next pandemic is to develop a quick-reaction force that would activate under prespecified criteria to manage reprioritization of all science and support activities to address pandemic response priorities at the velocity of relevance.

Chimeric Virus-like Particle-Based COVID-19 Vaccine Confers Strong Protection against SARS-CoV-2 Viremia in K18-hACE2 Mice.

Virus-like particles (VLPs) are highly immunogenic and versatile subunit vaccines composed of multimeric viral proteins that mimic the whole virus but lack genetic material. Due to the lack of infectivity, VLPs are being developed as safe and effective vaccines against various infectious diseases. In this study, we generated a chimeric VLP-based COVID-19 vaccine stably produced by HEK293T cells. The chimeric VLPs contain the influenza virus A matrix (M1) proteins and the SARS-CoV-2 Wuhan strain spike (S) proteins with a deletion of the polybasic furin cleavage motif and a replacement of the transmembrane and cytoplasmic tail with that of the influenza virus hemagglutinin (HA). These resulting chimeric S-M1 VLPs, displaying S and M1, were observed to be enveloped particles that are heterogeneous in shape and size. The intramuscular vaccination of BALB/c mice in a prime-boost regimen elicited high titers of S-specific IgG and neutralizing antibodies. After immunization and a challenge with SARS-CoV-2 in K18-hACE2 mice, the S-M1 VLP vaccination resulted in a drastic reduction in viremia, as well as a decreased viral load in the lungs and improved survival rates compared to the control mice. Balanced Th1 and Th2 responses of activated S-specific T-cells were observed. Moderate degrees of inflammation and viral RNA in the lungs and brains were observed in the vaccinated group; however, brain lesion scores were less than in the PBS control. Overall, we demonstrate the immunogenicity of a chimeric VLP-based COVID-19 vaccine which confers strong protection against SARS-CoV-2 viremia in mice.

A systematic analysis of ultrastructural lesions in the Plasmodium coatneyi splenectomized rhesus macaque model of severe malaria.

Plasmodium falciparum remains one of the world's deadliest diseases and with ongoing concerns of evolving drug resistance, there is a need for continued refinement of the Plasmodium coatneyi infection model in macaques to study severe malaria. As such, the systemic ultrastructural lesions associated with P. coatneyi infection in splenectomized rhesus macaques was evaluated in 6 animals. Autopsy samples from multiple areas of the central nervous system (CNS), kidneys, heart, liver, and lungs of all 6 animals were processed for electron microscopy. A systematic analysis of the ultrastructural changes associated with the plasmodium was undertaken by multiple pathologists to ensure consensus. All tissues exhibited marked sequestration of infected red blood cells comprised either of cytoadherence to endothelium or rosette formation, associated with variable degrees of host cell damage in a range of tissues that in severe cases resulted in necrosis. This is the first complete systemic evaluation of ultrastructural tissue lesions in P. coatneyi-infected rhesus macaques, and the findings have important implications evaluating of the use of this model for the study of severe malaria caused by P. falciparum in humans.

Limited and Short-Lasting Virus Neutralizing Titers Induced by Inactivated SARS-CoV-2 Vaccine.

In vitro determination of severe acute respiratory syndrome coronavirus 2 neutralizing antibodies induced in serum samples from recipients of the CoronaVac vaccine showed a short protection period against the original virus strain and limited protection against variants of concern. These data provide support for vaccine boosters, especially variants of concern circulate.

Combined Fatal Systemic Chlamydia sp. and Aeromonas sobria Infection in Juvenile Siamese Crocodiles ( Crocodylus siamensis).

Chlamydial infections in crocodiles have been described in several countries and in several different species. These are typically associated with severe pharyngitis and conjunctivitis, with death occurring secondary to compromise of the upper respiratory tract due to obstruction of the trachea. A population of ranched Siamese crocodiles in central Thailand experienced an epizootic of sudden death in juvenile animals. The affected animals had fulminant systemic disease primarily involving the liver and spleen but also affecting the kidneys, heart, and the whole of the respiratory tract. Chlamydia sp. were noted in liver and spleen during histopathological examination and confirmed with transmission electron microscopy and polymerase chain reaction (PCR). The sequence of the PCR product suggested a novel Chlamydia sp. of Siamese crocodiles. Crocodile farming represents an important economy in several parts of the world. Epizootics, such as the one described in this manuscript in association with Chlamydia sp., can have devastating impact on the industry and represent a potential zoonosis of significant public health concern. This is the first report of Chlamydia sp. and Aeromonas sobria causing systemic disease in crocodiles as well as the first histopathological and ultrastructural description of Chlamydia infection in Siamese crocodiles.

Characterization of the rhesus macaque (Macaca mulatta) scrub typhus model: Susceptibility to intradermal challenge with the human pathogen Orientia tsutsugamushi Karp.

BACKGROUND: Scrub typhus is an important endemic disease in tropical Asia caused by Orientia tsutsugamushi for which no effective broadly protective vaccine is available. The successful evaluation of vaccine candidates requires well-characterized animal models and a better understanding of the immune response against O. tsutsugamushi. While many animal species have been used to study host immunity and vaccine responses in scrub typhus, only limited data exists in non-human primate (NHP) models. METHODOLOGY/PRINCIPLE FINDINGS: In this study we evaluated a NHP scrub typhus disease model based on intradermal inoculation of O. tsutsugamushi Karp strain in rhesus macaques (n = 7). After an intradermal inoculation with 106 murine LD50 of O. tsutsugamushi at the anterior thigh (n = 4) or mock inoculum (n = 3), a series of time course investigations involving hematological, biochemical, molecular and immunological assays were performed, until day 28, when tissues were collected for pathology and immunohistochemistry. In all NHPs with O. tsutsugamushi inoculation, but not with mock inoculation, the development of a classic eschar with central necrosis, regional lymphadenopathy, and elevation of body temperature was observed on days 7-21 post inoculation (pi); bacteremia was detected by qPCR on days 6-18 pi; and alteration of liver enzyme function and increase of white blood cells on day 14 pi. Immune assays demonstrated raised serum levels of soluble cell adhesion molecules, anti-O. tsutsugamushi-specific antibody responses (IgM and IgG) and pathogen-specific cell-mediated immune responses in inoculated macaques. The qPCR assays detected O. tsutsugamushi in eschar, spleen, draining and non-draining lymph nodes, and immuno-double staining demonstrated intracellular O. tsutsugamushi in antigen presenting cells of eschars and lymph nodes. CONCLUSIONS/SIGNIFICANCE: These data show the potential of using rhesus macaques as a scrub typhus model, for evaluation of correlates of protection in both natural and vaccine induced immunity, and support the evaluation of future vaccine candidates against scrub typhus.

Strong interferon-gamma mediated cellular immunity to scrub typhus demonstrated using a novel whole cell antigen ELISpot assay in rhesus macaques and humans.

Scrub typhus is a febrile infection caused by the obligate intracellular bacterium Orientia tsutsugamushi, which causes significant morbidity and mortality across the Asia-Pacific region. The control of this vector-borne disease is challenging due to humans being dead-end hosts, vertical maintenance of the pathogen in the vector itself, and a potentially large rodent reservoir of unclear significance, coupled with a lack of accurate diagnostic tests. Development of an effective vaccine is highly desirable. This however requires better characterization of the natural immune response of this neglected but important disease. Here we implement a novel IFN-γ ELISpot assay as a tool for studying O. tsutsugamushi induced cellular immune responses in an experimental scrub typhus rhesus macaque model and human populations. Whole cell antigen for O. tsutsugamushi (OT-WCA) was prepared by heat inactivation of Karp-strain bacteria. Rhesus macaques were infected intradermally with O. tsutsugamushi. Freshly isolated peripheral blood mononuclear cells (PBMC) from infected (n = 10) and uninfected animals (n = 5) were stimulated with OT-WCA, and IFN-γ secreting cells quantitated by ELISpot assay at five time points over 28 days. PBMC were then assayed from people in a scrub typhus-endemic region of Thailand (n = 105) and responses compared to those from a partially exposed population in a non-endemic region (n = 14), and to a naïve population in UK (n = 12). Mean results at Day 0 prior to O. tsutsugamushi infection were 12 (95% CI 0-25) and 15 (2-27) spot-forming cells (SFC)/106 PBMC for infected and control macaques respectively. Strong O. tsutsugamushi-specific IFN-γ responses were seen post infection, with ELISpot responses 20-fold higher than baseline at Day 7 (mean 235, 95% CI 200-270 SFC/106 PBMC), 105-fold higher at Day 14 (mean 1261, 95% CI 1,097-1,425 SFC/106 PBMC), 125-fold higher at Day 21 (mean 1,498, 95% CI 1,496-1,500 SFC/106 PBMC) and 118-fold higher at Day 28 (mean 1,416, 95% CI 1,306-1,527 SFC/106 PBMC). No significant change was found in the control group at any time point compared to baseline. Humans from a scrub typhus endemic region of Thailand had mean responses of 189 (95% CI 88-290) SFC/106 PBMC compared to mean responses of 40 (95% CI 9-71) SFC/106 PBMC in people from a non-endemic region and 3 (95% CI 0-7) SFC/106 PBMC in naïve controls. In summary, this highly sensitive assay will enable field immunogenicity studies and further characterization of the host response to O. tsutsugamushi, and provides a link between human and animal models to accelerate vaccine development.

Accelerated senescence in skin in a murine model of radiation-induced multi-organ injury.

Accidental high-dose radiation exposures can lead to multi-organ injuries, including radiation dermatitis. The types of cellular damage leading to radiation dermatitis are not completely understood. To identify the cellular mechanisms that underlie radiation-induced skin injury in vivo, we evaluated the time-course of cellular effects of radiation (14, 16 or 17 Gy X-rays; 0.5 Gy/min) in the skin of C57BL/6 mice. Irradiation of 14 Gy induced mild inflammation, observed histologically, but no visible hair loss or erythema. However, 16 or 17 Gy radiation induced dry desquamation, erythema and mild ulceration, detectable within 14 days post-irradiation. Histological evaluation revealed inflammation with mast cell infiltration within 14 days. Fibrosis occurred 80 days following 17 Gy irradiation, with collagen deposition, admixed with neutrophilic dermatitis, and necrotic debris. We found that in cultures of normal human keratinocytes, exposure to 17.9 Gy irradiation caused the upregulation of p21/waf1, a marker of senescence. Using western blot analysis of 17.9 Gy-irradiated mice skin samples, we also detected a marker of accelerated senescence (p21/waf1) 7 days post-irradiation, and a marker of cellular apoptosis (activated caspase-3) at 30 days, both preceding histological evidence of inflammatory infiltrates. Immunohistochemistry revealed reduced epithelial stem cells from hair follicles 14-30 days post-irradiation. Furthermore, p21/waf1 expression was increased in the region of the hair follicle stem cells at 14 days post 17 Gy irradiation. These data indicate that radiation induces accelerated cellular senescence in the region of the stem cell population of the skin.

Nonlesions, misdiagnoses, missed diagnoses, and other interpretive challenges in fish histopathology studies: a guide for investigators, authors, reviewers, and readers.

Differentiating salient histopathologic changes from normal anatomic features or tissue artifacts can be decidedly challenging, especially for the novice fish pathologist. As a consequence, findings of questionable accuracy may be reported inadvertently, and the potential negative impacts of publishing inaccurate histopathologic interpretations are not always fully appreciated. The objectives of this article are to illustrate a number of specific morphologic findings in commonly examined fish tissues (e.g., gills, liver, kidney, and gonads) that are frequently either misdiagnosed or underdiagnosed, and to address related issues involving the interpretation of histopathologic data. To enhance the utility of this article as a guide, photomicrographs of normal and abnormal specimens are presented. General recommendations for generating and publishing results from histopathology studies are additionally provided. It is hoped that the furnished information will be a useful resource for manuscript generation, by helping authors, reviewers, and readers to critically assess fish histopathologic data.

Induction of rhabdomyosarcoma by embedded military-grade tungsten/nickel/cobalt not by tungsten/nickel/iron in the B6C3F1 mouse.

Continued improvements in the ballistic properties of military munitions have led to metal formulations for which little are known about the long-term health effects. Previously we have shown that a military-grade tungsten alloy comprised of tungsten, nickel, and cobalt, when embedded into the leg muscle of F344 rats to simulate a fragment wound, induces highly aggressive metastatic rhabdomyosarcomas. An important follow-up when assessing a compound's carcinogenic potential is to test it in a second rodent species. In this study, we assessed the health effects of embedded fragments of 2 military-grade tungsten alloys, tungsten/nickel/cobalt and tungsten/nickel/iron, in the B6C3F1 mouse. Implantation of tungsten/nickel/cobalt pellets into the quadriceps muscle resulted in the formation of a rhabdomyosarcoma around the pellet. Conversely, implantation of tungsten/nickel/iron did not result in tumor formation. Unlike what was seen in the rat model, the tumors induced by the tungsten/nickel/cobalt did not exhibit aggressive growth patterns and did not metastasize.

Laparoscopic colpotomy using monopolar electrocautery: does power really matter?

OBJECTIVE: The purpose of this study was to assess the extent and rate of vaginal tissue injury associated with the utilization of various monopolar electrosurgical power settings when laparoscopically transecting vaginal tissue. METHODS: This is an Institutional Animal Care and Use Committee-approved prospective, paired, single-blinded study. Externalized porcine vagina was transected using monopolar energy at 30, 50, and 80 W in the cut mode with laparoscopic Endo Shears. The slides were prepared and stained with both hematoxylin-eosin and Masson trichrome and were examined by board-certified veterinary pathologists blinded to the study. RESULTS: There were 18 swine; each animal was tested on all 3 power settings (n = 54). Tissue injury was measured to a mean (SD) of 767 (519) µm at 30 W, 690 (600) µm at 50 W, and 556 (470) µm at 80 W. When comparing the monopolar settings, the results were as follows: 30 versus 50 W (P = 0.33), 30 versus 80 W (P = 0.067), and 50 versus 80 W (P = 0.17). The mean (SD) time for complete transection was measured at each power setting (n = 18), with 35.8 (5.4) seconds for 30 W, 13.5 (5.5) seconds for 50 W, and 8.4 (5.1) seconds for 80 W (P < 0.001). There was a statistically significant difference in the mean (SD) rates of injury, with 20.8 (8.8) µm/s at 30 W, 39.8 (11.8) µm/s at 50 W, and 50.1 (19.2) µm/s at 80 W (P = 0.01). CONCLUSIONS: Using various power settings of monopolar energy may not make a significant difference in swine vaginal tissue damage at the time of colpotomy. However, there was a significant difference in the times and rates at which tissue was transected when using higher powers. We recommend using the 50- or 80-W setting, as this will likely decrease surgical times without altering vaginal tissue damage.

The Role of the Component Metals in the Toxicity of Military-Grade Tungsten Alloy.

Tungsten-based composites have been recommended as a suitable replacement for depleted uranium. Unfortunately, one of these mixtures composed of tungsten (W), nickel (Ni) and cobalt (Co) induced rhabdomyosarcomas when implanted into the leg muscle of laboratory rats and mice to simulate a shrapnel wound. The question arose as to whether the neoplastic effect of the mixture could be solely attributed to one or more of the metal components. To investigate this possibility, pellets with one or two of the component metals replaced with an identical amount of the biologically-inert metal tantalum (Ta) were manufactured and implanted into the quadriceps of B6C3F1 mice. The mice were followed for two years to assess potential adverse health effects. Implantation with WTa, CoTa or WNiTa resulted in decreased survival, but not to the level reported for WNiCo. Sarcomas in the implanted muscle were found in 20% of the CoTa-implanted mice and 5% of the WTa- and WCoTa-implanted rats and mice, far below the 80% reported for WNiCo-implanted mice. The data obtained from this study suggested that no single metal is solely responsible for the neoplastic effects of WNiCo and that a synergistic effect of the three metals in tumor development was likely.

Costal mesenchymal chondrosarcoma with diffuse pleural and pericardial explantation in a pygmy goat.

A 3 year old intact male pygmy goat developed progressive weakness and eventual recumbancy over the course of 1 week, while maintaining its ability to eat and drink. The animal died and at necropsy, the parietal pleural surfaces and the pericardial surface were noted to be covered with firm, white, variably sized nodules that often formed linear arrays or coalesced into larger clumped aggregates. The visceral pleural surfaces of the ventral lung lobes were also covered with similar nodules. Histopathological and immunohistochemical evaluation of the submitted tissues revealed a diagnosis of mesenchymal chondrosarcoma with extensive seeding throughout the thoracic cavity.

Mammary carcinoma in a male rhesus macaque (Macaca mulatta): histopathology and immunohistochemistry of ductal carcinoma in situ.

BACKGROUND: A mammary nodule was noted in a male rhesus macaque during physical examination. METHODS AND RESULTS: Histopathological and immunohistochemical analysis was performed. Ductal carcinoma in situ was confirmed. CONCLUSIONS: To date, there are two reports of mammary carcinoma in male non-human primates, and none in the rhesus macaque.

Accelerated hematopoietic syndrome after radiation doses bridging hematopoietic (H-ARS) and gastrointestinal (GI-ARS) acute radiation syndrome: early hematological changes and systemic inflammatory response syndrome in minipig.

PURPOSE: To characterize acute radiation syndrome (ARS) sequelae at doses intermediate between the bone marrow (H-ARS) and full gastrointestinal (GI-ARS) syndrome. METHODS: Male minipigs, approximately 5 months old, 9-12 kg in weight, were irradiated with Cobalt-60 (total body, bilateral gamma irradiation, 0.6 Gy/min). Endpoints were 10-day survival, gastrointestinal histology, plasma citrulline, bacterial translocation, vomiting, diarrhea, vital signs, systemic inflammatory response syndrome (SIRS), febrile neutropenia (FN). RESULTS: We exposed animals to doses (2.2-5.0 Gy) above those causing H-ARS (1.6-2.0 Gy), and evaluated development of ARS. Compared to what was observed during H-ARS (historical data: Moroni et al. 2011a , 2011c ), doses above 2 Gy produced signs of increasingly severe pulmonary damage, faster deterioration of clinical conditions, and faster increases in levels of C-reactive protein (CRP). In the range of 4.6-5.0 Gy, animals died by day 9-10; signs of the classic GI syndrome, as measured by diarrhea, vomiting and bacterial translocation, did not occur. At doses above 2 Gy we observed transient reduction in circulating citrulline levels, and animals exhibited earlier depletion of blood elements and faster onset of SIRS and FN. CONCLUSIONS: An accelerated hematopoietic subsyndrome (AH-ARS) is observed at radiation doses between those producing H-ARS and GI-ARS. It is characterized by early onset of SIRS and FN, and greater lung damage, compared to H-ARS.

Esophageal adenocarcinoma in a 53-year-old mata mata turtle (Chelus fimbriatus).

A 53-yr-old male captive mata mata turtle (Chelus fimbriatus) was examined following sudden death. The animal was in good nutritional, muscular and postmortem condition. The esophageal wall was circumferentially expanded by a discrete, oblong, irregular, tan, and soft contiguous glandlike structure. Histologically, the mass comprised uneven, sometimes cavitated islands of polygonal neoplastic cells consistent with an esophageal adenocarcinoma. In addition, peripheral to the mass, there was glandular epithelial hyperplasia, dysplasia, and multifocal heterophilic and lymphohistiocytic adenitis. Neoplastic cells expressed pancytokeratins; however, they demonstrated no immunoreactivity to vimentin, chromogranin, synaptophysin, and thyroglobulin. Additional findings included multifocal to coalescing areas of cortical fibrosis and membranous glomerulonephritis affecting both kidneys, and a focal hepatocellular adenoma.

The Gottingen minipig is a model of the hematopoietic acute radiation syndrome: G-colony stimulating factor stimulates hematopoiesis and enhances survival from lethal total-body γ-irradiation.

PURPOSE: We are characterizing the Gottingen minipig as an additional large animal model for advanced drug testing for the acute radiation syndrome (ARS) to enhance the discovery and development of novel radiation countermeasures. Among the advantages provided by this model, the similarities to human hematologic parameters and dynamics of cell loss/recovery after irradiation provide a convenient means to compare the efficacy of drugs known to affect bone marrow cellularity and hematopoiesis. METHODS AND MATERIALS: Male Gottingen minipigs, 4 to 5 months old and weighing 9 to 11 kg, were used for this study. We tested the standard off-label treatment for ARS, rhG-CSF (Neupogen, 10 µg/kg/day for 17 days), at the estimated LD70/30 total-body γ-irradiation (TBI) radiation dose for the hematopoietic syndrome, starting 24 hours after irradiation. RESULTS: The results indicated that granulocyte colony stimulating factor (G-CSF) enhanced survival, stimulated recovery from neutropenia, and induced mobilization of hematopoietic progenitor cells. In addition, the administration of G-CSF resulted in maturation of monocytes/macrophages. CONCLUSIONS: These results support continuing efforts toward validation of the minipig as a large animal model for advanced testing of radiation countermeasures and characterization of the pathophysiology of ARS, and they suggest that the efficacy of G-CSF in improving survival after total body irradiation may involve mechanisms other than increasing the numbers of circulating granulocytes.

Ciprofloxacin modulates cytokine/chemokine profile in serum, improves bone marrow repopulation, and limits apoptosis and autophagy in ileum after whole body ionizing irradiation combined with skin-wound trauma.

Radiation combined injury (CI) is a radiation injury (RI) combined with other types of injury, which generally leads to greater mortality than RI alone. A spectrum of specific, time-dependent pathophysiological changes is associated with CI. Of these changes, the massive release of pro-inflammatory cytokines, severe hematopoietic and gastrointestinal losses and bacterial sepsis are important treatment targets to improve survival. Ciprofloxacin (CIP) is known to have immunomodulatory effect besides the antimicrobial activity. The present study reports that CIP ameliorated pathophysiological changes unique to CI that later led to major mortality. B6D2F1/J mice received CI on day 0, by RI followed by wound trauma, and were treated with CIP (90 mg/kg p.o., q.d. within 2 h after CI through day 10). At day 10, CIP treatment not only significantly reduced pro-inflammatory cytokine and chemokine concentrations, including interleukin-6 (IL-6) and KC (i.e., IL-8 in human), but it also enhanced IL-3 production compared to vehicle-treated controls. Mice treated with CIP displayed a greater repopulation of bone marrow cells. CIP also limited CI-induced apoptosis and autophagy in ileal villi, systemic bacterial infection, and IgA production. CIP treatment led to LD(0/10) compared to LD(20/10) for vehicle-treated group after CI. Given the multiple beneficial activities of CIP shown in our experiments, CIP may prove to be a useful therapeutic drug for CI.

Abnormal microRNA expression in Ts65Dn hippocampus and whole blood: contributions to Down syndrome phenotypes.

Down syndrome (DS; trisomy 21) is one of the most common genetic causes of intellectual disability, which is attributed to triplication of genes located on chromosome 21. Elevated levels of several microRNAs (miRNAs) located on chromosome 21 have been reported in human DS heart and brain tissues. The Ts65Dn mouse model is the most investigated DS model with a triplicated segment of mouse chromosome 16 harboring genes orthologous to those on human chromosome 21. Using ABI TaqMan miRNA arrays, we found a set of miRNAs that were significantly up- or downregulated in the Ts65Dn hippocampus compared to euploid controls. Furthermore, miR-155 and miR-802 showed significant overexpression in the Ts65Dn hippocampus, thereby confirming results of previous studies. Interestingly, miR-155 and miR-802 were also overexpressed in the Ts65Dn whole blood but not in lung tissue. We also found overexpression of the miR-155 precursors, pri- and pre-miR-155 derived from the miR-155 host gene, known as B cell integration cluster, suggesting enhanced biogenesis of miR-155. Bioinformatic analysis revealed that neurodevelopment, differentiation of neuroglia, apoptosis, cell cycle, and signaling pathways including ERK/MAPK, protein kinase C, phosphatidylinositol 3-kinase, m-TOR and calcium signaling are likely targets of these miRNAs. We selected some of these potential gene targets and found downregulation of mRNA encoding Ship1, Mecp2 and Ezh2 in Ts65Dn hippocampus. Interestingly, the miR-155 target gene Ship1 (inositol phosphatase) was also downregulated in Ts65Dn whole blood but not in lung tissue. Our findings provide insights into miRNA-mediated gene regulation in Ts65Dn mice and their potential contribution to impaired hippocampal synaptic plasticity and neurogenesis, as well as hemopoietic abnormalities observed in DS.

Laparoscopic hysterectomy using various energy sources in swine: a histopathologic assessment.

OBJECTIVE: Analyze energy-induced damage to the swine vagina during laparoscopic hysterectomy. STUDY DESIGN: Laparoscopic colpotomy was performed in swine using ultrasonic, monopolar, and bipolar energy. Specimens (n = 22) from 13 swine were stained with hematoxylin and eosin and Masson's trichrome for energy-related damage. The distal scalpel-cut margin was used as reference. Energy induced damage was assessed by gynecologic and veterinary pathologists blinded to energy source. RESULTS: Injury was most apparent on Masson's trichrome, demonstrating clear injury demarcation, allowing consistent, quantitative damage measurements. Mean injury was 0 ± 0 µM (scalpel, n = 22), 782 ± 359 µM (ultrasonic, n = 7), 2016 ± 1423 µM (monopolar, n = 8), and 3011 ± 1239 µM (bipolar, n = 7). Using scalpel as the reference, all were significant (P < .001). CONCLUSION: All energy sources demonstrated tissue damage, with ultrasonic showing the least and bipolar the greatest. Further study of tissue damage relative to cuff closure at laparoscopic hysterectomy is warranted.