Optical Dynamic Analysis of Thrombus Inside a Centrifugal Blood Pump During Extracorporeal Mechanical Circulatory Support in a Porcine Model.

Complications due to pump thrombus remain the weak point of mechanical circulatory support (MCS), such as the use of a left ventricular assist device (LVAD) or extracorporeal membrane oxygenation, leading to poor outcomes. Hyperspectral imaging (HSI) is an effective imaging method using a hyperspectral (HS) camera, which comprises a spectrophotometer and a charge-coupled device camera to discriminate thrombus from whole blood. Animal experiments were conducted to analyze dynamic imaging of thrombus inside a prototype of a hydrodynamically levitated centrifugal blood pump using an HSI system. Six pigs were divided into a venous circulation group (n = 3) and an arterial circulation group (n = 3). Inflow and outflow cannulae were inserted into the jugular veins in the venous circulation group. The latter simulated an LVAD application. To create thrombogenic conditions, pump flow was maintained at 1 L/min without anticoagulation. An image of the bottom surface of the pump was captured by the HS camera every 4 nm over the wavelength range of 608-752 nm. Real-time dynamic images of the inside of the pump were displayed on the monitor. Appearance of an area displaying thrombus was detected within 24 h after the start of the circulation in every experiment. This imaging system also succeeded in determining the origins of pump thrombus: from inside the pump in two cases, and from outside in four cases. Two main possible sources of pump thrombus originating outside the pump were identified on autopsy: wedge thrombus around the inflow cannula; and string-like thrombus at the junction between the pump inlet and circuit tube. The results of this study from close observation of the changing appearance of pump thrombus may contribute to improvements in the safety of extracorporeal MCS.

[A case of serous surface papillary carcinoma of the peritoneum metastatic to the brain].

A case of brain metastasis from peritoneal serous surface papillary carcinoma (SSPC) was reported. This 68-year-old woman was admitted to our department because of decreased consciousness level for the last few days. Her medical past history revealed breast cancer and SSPC of the peritoneum at the age of 64. On admission she was comatose and irritable. Serum and urine examination revealed the syndrome of inappropriate secretion of antidiuretic hormone that was treated with strict restriction of water intake. MRI of the brain demonstrated a nonspecific mass in the subcortical area of the right superior parietal lobule. The mass was successfully removed in en bloc fashion. Pathological diagnosis was SSPC that was compatible with the previous diagnosis obtained from the peritoneum four years previously. Although the patient received whole brain radiation therapy postoperatively, her condition deteriorated rapidly. She died four months after brain surgery. SSPC of the peritoneum is a rare malignant tumor that is defined as a primary tumor histologically indistinguishable from serous carcinoma of the ovary, diffusely involving the peritoneal surface but sparing or only superficially invading the ovaries. Because of the prolongation of survival resulting from advanced chemotherapy for SSPC of the peritoneum, more patients live long enough to develop brain metastasis. Therefore, SSPC of the peritoneum should be kept in mind in the differential diagnosis of a primary site for brain metastasis.

Transplantation of neuronal cells induced from human mesenchymal stem cells improves neurological functions after stroke without cell fusion.

The options for treating stroke are limited, but stem cells hold promise as a therapy because of their multipotency. Neuronal cells derived from mesenchymal stem cells (MSC) were reported to have more therapeutic effect than MSCs. For elucidating the therapeutic mechanism of neuronal cells, here we generated a model of focal cerebral infarction by performing left common carotid artery occlusion in adult gerbils. We transfected human trabecular bone-derived MSCs (hMSCs) with the Notch intracellular domain to induce their differentiation into neuronal cells (hN-MSCs). These cells were stereotaxically transplanted into the local ischemic hemisphere 4 days after the occlusion. Behavioral analyses were conducted 28 days after transplantation, and then fluorescence in situ hybridization (FISH) and a histological evaluation were performed. Histologically, transplanted cells were distributed around the periinfarct region, and approximately 8.5% and 4.2% of hN-MSCs and hMSCs survived, respectively; 53.2% ± 9.6% of hN-MSCs were microtubule-associated protein 2(+) (MAP-2(+) ) and extended neurites, whereas only 0.9% ± 0.3% of hMSCs were MAP-2(+) . In FISH, human nucleus-specific signals were detected in both hN-MSCs and hMSCs grafted brains, but no transplanted cell had a merged gerbil-specific nuclear signals. hN-MSC-transplanted animals showed significantly better recovery than animals given control vehicle in the T-maze, bilateral asymmetry, and open field tests. These findings suggested that hN-MSCs have greater therapeutic potential than hMSCs for stroke and that cell fusion does not primarily contribute to the therapeutic mechanism of MSC transplantation.

Two region-dependent pathways of eosinophilic neuronal death after transient cerebral ischemia.

Various types of eosinophilic neurons (ENs) are found in the post-ischemic brain. We examined the temporal profile of ENs in the core and peripheral regions of the ischemic cortex, and analyzed the relationship to the expression of various cell death-related factors. Unilateral forebrain ischemia was induced in Mongolian gerbils by transient common carotid artery occlusions, and the brains from 3 h to 2 weeks post-ischemia were prepared for morphometric and immunohistochemical analysis of ENs. ENs with minimally abnormal nuclei and swollen cell bodies appeared at 3 h in the ischemic core and at 12 h in the periphery. In both locations multiple cell death-related factors including calcium, micro-calpain, cathepsin D, 78 kDa glucose-regulated protein (GRP78) and ubiquitin were activated. In the ischemic core, pyknosis and irregularly atrophic cytoplasm peaked at 12 h, which was associated with significant increases in staining for calcium and micro-calpain. ENs with pyknosis and scant cytoplasm peaked at 4 days and were positive for TUNEL and calcium staining. In the ischemic periphery, ENs had slightly atrophic cytoplasm and sequentially developed pyknosis, karyorrhexis and karyolysis over 1 week. These cells were positive for TUNEL and calcium staining. All types of EN were negative for caspase 3. There may be two region-dependent pathways of EN changes in the post-ischemic brain: pyknosis with cytoplasmic shrinkage in the core, and nuclear disintegration with slightly atrophic cytoplasm in the periphery. This difference coordinates different activation patterns of cell death-related factors in ENs.

Transient cognitive deficits are associated with the reversible accumulation of amyloid precursor protein after mild traumatic brain injury.

Mild traumatic brain injury (MTBI) may frequently cause transient behavioral abnormalities without observable morphological findings. In this study, we investigated neuropathological mechanisms underlying transient cognitive deficits after MTBI. Mongolian gerbils were subjected to experimental MTBI. At various time points after injury, behavioral changes were evaluated by the open-field test and T-maze test, and immunohistochemistry of microtubule-associated protein (MAP2) and amyloid precursor protein (APP) was performed to examine disruptions of the neuronal cytoskeleton and axonal transport, respectively. Transient cognitive deficits were observed after MTBI. Sustained MAP2 loss was found within the cortical impact site, but not the hippocampus. Transient APP accumulation at the same time as transient cognitive deficits occurred in the ipsilateral hemisphere, particularly in the subcortical white matter. These results suggest that the axonal dysfunction indicated by the reversible APP accumulation in the white matter, but not the sustained neuronal cytoskeletal damage reflected by the cortical MAP2 loss confined to the impact site, is responsible for the transient functional deficits after MTBI.

Heterogeneous hyperactivity and distribution of ischemic lesions after focal cerebral ischemia in Mongolian gerbils.

Various types of poststroke hyperactivity exist in humans, but studies of each mechanism using animal models are scarce. We aimed to analyze the heterogeneity of postischemic hyperlocomotion and to identify the ischemic lesions responsible for postischemic hyperlocomotion in rodent models of focal ischemia. Mongolian gerbils underwent right common carotid artery occlusion (CCAO) for 10 or 20 min. At 24 h, 2 days, and 7 days postischemia, we performed quantitative and qualitative locomotor analysis and correlated these results with the extent of ischemic lesions. Intermittent explosive hyperlocomotion was induced transiently in a 10-min CCAO group at 24 h after ischemia and continual unexplosive hyperlocomotion persisted for 7 days in the 20-min CCAO animals. Selective neuronal death, confined to the hippocampal cornu ammonis 1 (CA1), was observed in the 10-min CCAO group and widespread cortical and basal ganglia infarction was observed in the 20-min CCAO group. Amyloid precursor protein was transiently observed in the hippocampus at 24 h postischemia in the 10-min CCAO animals, while it was widely distributed over the ischemic regions throughout the 7 days postischemia in the 20-min CCAO animals. Incidence maps and correlation analysis revealed hippocampal neuronal death of the CA1 sector and widespread hemispheric infarction, including the cortex, as the region responsible for the 10-min and 20-min CCAO-induced hyperactivity, respectively. Two distinct types of locomotor hyperactivity were observed that varied with regard to the distribution of the ischemic lesion, that is, hippocampal neuronal death and widespread infarction involving the cortex. These two types of locomotor hyperactivity appear to be models of the different types of poststroke hyperactivity seen in stroke patients.

Transient versus prolonged hyperlocomotion following lateral fluid percussion injury in mongolian gerbils.

Posttraumatic hyperactivity is a neurobehavioral symptom commonly seen in patients after traumatic brain injury (TBI). No useful animal model has yet been established for evaluation of this important symptom. We induced either mild (MILD, 0.7-0.9 atm) or moderate (MOD, 1.3-1.6 atm) lateral fluid percussion injury (LFPI) in Mongolian gerbils. Open-field and T-maze tests were used during a 7-day period after the trauma. All animals were perfusion fixed for histopathological examinations. Transient locomotor hyperactivity was found with a peak at 6 hr after injury in the MILD animals, whereas MOD animals showed prolonged and severe hyperlocomotion throughout the 7-day posttrauma period (P < 0.0001). Interestingly, the temporal profile of the posttraumatic hyperactivity was similar to that of the working memory deficit in both injury groups. Histological examination revealed significant neural tissue damages, including cortical necrosis, white matter rarefaction, and neuronal loss in the hippocampus in the ipsilateral hemisphere of the MOD animals, vs. only negligible changes in the MILD animals. Correlation analysis revealed that the volume of white matter lesions was significantly correlated with both posttraumatic hyperactivity (r = 0.591, P < 0.01) and working memory deficit (r = -0.859, P < 0.0001). Taken together, our findings confirm the successful reproduction of posttraumatic hyperactivity following experimental TBI. The posttraumatic hyperlocomotion probably shared pathomechanisms common to those of cognitive dysfunction caused by LFPI, supporting the speculation from previous studies that some neurobehavioral abnormities intimately correlate with TBI-induced cognitive dysfunction. Histopathologically, significant involvement of white matter damage in the posttraumatic functional deficits was indicated.

Transition of areas of eosinophilic neurons and reactive astrocytes to delayed cortical infarcts after transient unilateral forebrain ischemia in Mongolian gerbils.

The fate of postischemic tissues containing eosinophilic neurons (ENs), whether they remain viable or evolve into infarction, is largely unknown. We analyzed the time profile and distribution of ENs, reactive astrocytes (RAs), and infarction after transient cerebral ischemia. Unilateral forebrain ischemia was induced in Mongolian gerbils by two 10-min unilateral common carotid artery occlusions with a 5-h interval, and the brains at 24 h, 4 days, and 2, 4, and 16 weeks were prepared for morphometric analysis. Intra-ischemic laser Doppler flowmetry revealed significant ischemia, deeper in the anterior cortex, during carotid occlusion. Here, ENs appeared in the middle and deep layers at 24 h postischemia, and EN areas had extended to all cortical layers by 4 days. Large areas of high EN density turned into infarcts between 4 days and 4 weeks. In the posterior cortex, middle and deep cortical layers evolved low EN density areas without subsequent transformation into infarcts. RAs were consistently observed in areas with ENs, and RA areas with high EN density were largely transformed into infarcts between 4 days and 4 weeks postischemia. Areas of high, but not low, EN density were slowly transformed into infarcts after transient cerebral ischemia. Delayed astrocytic death took place in the RA areas with high EN density. In conclusion, density of ENs is an important indicator of delayed astrocytic death and infarction in postischemic tissue.

Helicobacter pylori is a fragile bacteria when stored at low and ultra-low temperatures.

BACKGROUND AND AIM: Usually, bacteria are cryopreserved for short-term storage at low and ultra-low temperatures. There are no reports as to whether Helicobacter pylori is a fragile bacteria when stored at low and ultra-low temperatures as compared with other intestinal bacteria. A study was done on seven H. pylori strains and other intestinal bacteria to compare different temperatures for storage of organisms in saline solution. METHODS: Seven H. pylori strains, specifically American Type Culture Collection (ATCC) strains 43504 and TN2GF4, and five strains isolated from the present patients were grown on a modified Skirrow's agar for H. pylori. Escherichia coli and Bacteroides distasonis, both representing isolates from the present patients, were grown on trypticase soy blood agar for E. coli, and EG agar for B. distasonis. Culture was for 4-5 days under microaerobic, aerobic and anaerobic conditions at 37 degrees C. Cells were harvested by scraping growth from the solid medium and into sterile saline. The cells were adjusted to concentrations of 109 viable cells/mL in saline and preserved at 4 degrees C, -20 degrees C, or -80 degrees C for 3 weeks before reculture under microaerobic, aerobic and anaerobic conditions at 37 degrees C for 7 days. After incubation, morphologically distinct colonies were counted, isolated, and identified by standard bacteriologic techniques. The H. pylori were morphologically analyzed by electronic microscopy before and after preservation. Mongolian gerbils were inoculated with the cryopreserved H. pylori to evaluate the bacterial infectivity. RESULTS: Six of the seven H. pylori strains failed to culture after being preserved at 4 degrees C, -20 degrees C, or -80 degrees C. Only ATCC 43504 could be cultured after freezing at -80 degrees C. The number of H. pylori ATCC 43504 before preservation was 9.0 +/- 0.5 (log10 no. organisms/mL) and decreased to 5.7 +/- 0.6 after preservation. Morphologically, all H. pylori except ATCC 43504 strains transformed from a bacillary to a coccoid form after preservation. In addition, none of the H. pylori strains could infect Mongolian gerbils after preservation. Escherichia coli and B. distasonis were recovered. Titers before and after 4 degrees C, -20 degrees C, and -80 degrees C, respectively, were 9.1 +/- 0.2, 8.9 +/- 0.5, 8.6 +/- 0.3, and 8.7 +/- 0.3 for E. coli and 9.1 +/- 0.4, 8.7 +/- 0.6, 8.6 +/- 0.5, and 8.8 +/- 0.3 for B. distasonis. CONCLUSIONS: Helicobacter pylori is a fragile bacteria for storage at low and ultra-low temperatures in comparison with other intestinal bacteria.

Neurological dysfunctions versus regional infarction volume after focal ischemia in Mongolian gerbils.

BACKGROUND AND PURPOSE: With advances in the therapy of stroke at the postacute phase, the use of animal models for chronological and region-specific evaluation of neurological function has become increasingly important. Our aim was to test long-term behavioral dysfunction in gerbils after focal ischemia and to correlate the results with the regional distribution of infarction in the coordinating cortical regions. METHODS: Repetitive unilateral hemispheric ischemia (two 10-minute occlusions, 5-hour interval) was induced in Mongolian gerbils. The elevated body swing test (EBST), bilateral asymmetry test (BAT), and T-maze test were performed to assess asymmetrical motor behavior, somatosensory deficit, and spatial cognitive dysfunction during 4 weeks after ischemia. The results were correlated against the regional infarction volume of the primary motor, somatosensory, and primary visual cortices at 4 weeks after ischemia. RESULTS: In all postischemic gerbils, persistent sensorimotor and cognitive dysfunctions were detectable throughout the postischemic period. Histological examination revealed that a cortical zone of infarction surrounded the selective neuronal death in the ipsilateral cerebral hemisphere. The regional infarction volumes of the primary motor, somatosensory, and visual cortices were significantly correlated with the scores of the EBST, BAT, and T-maze test, respectively. These combinations had the highest regression coefficient of all pairs. CONCLUSIONS: Postischemic motor and somatosensory functions were significantly correlated with regional infarction volumes in the corresponding cortical regions. In gerbils, visual abnormality could be independently detected by the T-maze test. Such regional analyses of ischemic lesions would be useful for investigating the functional outcomes of stroke therapy.