Intra-Arterial Transplantation of Allogeneic Mesenchymal Stem Cells Mounts Neuroprotective Effects in a Transient Ischemic Stroke Model in Rats: Analyses of Therapeutic Time Window and Its Mechanisms

Objective

Intra-arterial stem cell transplantation exerts neuroprotective effects for ischemic stroke. However, the optimal therapeutic time window and mechanisms have not been completely understood. In this study, we investigated the relationship between the timing of intra-arterial transplantation of allogeneic mesenchymal stem cells (MSCs) in ischemic stroke model in rats and its efficacy in acute phase.

Methods

Adult male Wistar rats weighing 200 to 250g received right middle cerebral artery occlusion (MCAO) for 90 minutes. MSCs (1×10⁶cells/ 1ml PBS) were intra-arterially injected at either 1, 6, 24, or 48 hours (1, 6, 24, 48h group) after MCAO. PBS (1ml) was intra-arterially injected to control rats at 1 hour after MCAO. Behavioral test was performed immediately after reperfusion, and at 3, 7 days after MCAO using the Modified Neurological Severity Score (mNSS). Rats were euthanized at 7 days after MCAO for evaluation of infarct volumes and the migration of MSCs. In order to explore potential mechanisms of action, the upregulation of neurotrophic factor and chemotactic cytokine (bFGF, SDF-1α) induced by cell transplantation was examined in another cohort of rats that received intra-arterial transplantation at 24 hours after recanalization then euthanized at 7 days after MCAO for protein assays.

Results

Behavioral test at 3 and 7 days after transplantation revealed that stroke rats in 24h group displayed the most robust significant improvements in mNSS compared to stroke rats in all other groups (p’s<0.05). Similarly, the infarct volumes of stroke rats in 24h group were much significantly decreased compared to those in all other groups (p’s<0.05). These observed behavioral and histological effects were accompanied by MSC survival and migration, with the highest number of integrated MSCs detected in the 24h group. Moreover, bFGF and SDF-1α levels of the infarcted cortex were highly elevated in the 24h group compared to control group (p’s<0.05).

Conclusions

These results suggest that intra-arterial allogeneic transplantation of MSCs provides post-stroke functional recovery and reduction of infarct volumes in ischemic stroke model of rats. The upregulation of bFGF and SDF-1α likely played a key mechanistic role in enabling MSC to afford functional effects in stroke. MSC transplantation at 24 hours after recanalization appears to be the optimal timing for ischemic stroke model, which should guide the design of clinical trials of cell transplantation for stroke patients.     

Angiographic Lesion Complexity Score and In-Hospital Outcomes after Percutaneous Coronary Intervention

Objective

We devised a percutaneous coronary intervention (PCI) scoring system based on angiographic lesion complexity and assessed its association with in-hospital complications.

Background

Although PCI is finding increasing application in patients with coronary artery disease, lesion complexity can lead to in-hospital complications.

Methods

Data from 3692 PCI patients were scored based on lesion complexity, defined by bifurcation, chronic total occlusion, type C, and left main lesion, along with acute thrombus in the presence of ST-segment elevation myocardial infarction (1 point assigned for each variable).

Results

The patients’ mean age was 67.5 +/- 10.8 years; 79.8% were male. About half of the patients (50.3%) presented with an acute coronary syndrome, and 2218 (60.1%) underwent PCI for at least one complex lesion. The patients in the higher-risk score groups were older (p < 0.001) and had present or previous heart failure (p = 0.02 and p = 0.01, respectively). Higher-risk score groups had significantly higher in-hospital event rates for death, heart failure, and cardiogenic shock (from 0 to 4 risk score; 1.7%, 4.5%, 6.3%, 7.1%, 40%, p < 0.001); bleeding with a hemoglobin decrease of >3.0 g/dL (3.1%, 11.0%, 13.1%, 10.3%, 28.6%, p < 0.001); and postoperative myocardial infarction (1.5%, 3.1%, 3.8%, 3.8%, 10%, p = 0.004), respectively. The association with adverse outcomes persisted after adjustment for known clinical predictors (odds ratio 1.72, p < 0.001).

Conclusion

The complexity score was cumulatively associated with in-hospital mortality and complication rate and could be used for event prediction in PCI patients.     

Medical Decision-Making Incapacity among Newly Diagnosed Older Patients with Hematological Malignancy Receiving First Line Chemotherapy: A Cross-Sectional Study of Patients and Physicians

Background

Decision-making capacity to provide informed consent regarding treatment is essential among cancer patients. The purpose of this study was to identify the frequency of decision-making incapacity among newly diagnosed older patients with hematological malignancy receiving first-line chemotherapy, to examine factors associated with incapacity and assess physicians’ perceptions of patients’ decision-making incapacity.

Methods

Consecutive patients aged 65 years or over with a primary diagnosis of malignant lymphoma or multiple myeloma were recruited. Decision-making capacity was assessed using the Structured Interview for Competency and Incompetency Assessment Testing and Ranking Inventory-Revised (SICIATRI-R). Cognitive impairment, depressive condition and other possible associated factors were also evaluated.

Results

Among 139 eligible patients registered for this study, 114 completed the survey. Of these, 28 (25%, 95% confidence interval [CI]: 17%-32%) were judged as having some extent of decision-making incompetency according to SICIATRI-R. Higher levels of cognitive impairment and increasing age were significantly associated with decision-making incapacity. Physicians experienced difficulty performing competency assessment (Cohen’s kappa -0.54).

Conclusions

Decision-making incapacity was found to be a common and under-recognized problem in older patients with cancer. Age and assessment of cognitive impairment may provide the opportunity to find patients that are at a high risk of showing decision-making incapacity.     

Suppression of Hydroxycinnamate Network Formation in Cell Walls of Rice Shoots Grown under Microgravity Conditions in Space

Network structures created by hydroxycinnamate cross-links within the cell wall architecture of gramineous plants make the cell wall resistant to the gravitational force of the earth. In this study, the effects of microgravity on the formation of cell wall-bound hydroxycinnamates were examined using etiolated rice shoots simultaneously grown under artificial 1 g and microgravity conditions in the Cell Biology Experiment Facility on the International Space Station. Measurement of the mechanical properties of cell walls showed that shoot cell walls became stiff during the growth period and that microgravity suppressed this stiffening. Amounts of cell wall polysaccharides, cell wall-bound phenolic acids, and lignin in rice shoots increased as the shoot grew. Microgravity did not influence changes in the amounts of cell wall polysaccharides or phenolic acid monomers such as ferulic acid (FA) and p-coumaric acid, but it suppressed increases in diferulic acid (DFA) isomers and lignin. Activities of the enzymes phenylalanine ammonia-lyase (PAL) and cell wall-bound peroxidase (CW-PRX) in shoots also increased as the shoot grew. PAL activity in microgravity-grown shoots was almost comparable to that in artificial 1 g-grown shoots, while CW-PRX activity increased less in microgravity-grown shoots than in artificial 1 g-grown shoots. Furthermore, the increases in expression levels of some class III peroxidase genes were reduced under microgravity conditions. These results suggest that a microgravity environment modifies the expression levels of certain class III peroxidase genes in rice shoots, that the resultant reduction of CW-PRX activity may be involved in suppressing DFA formation and lignin polymerization, and that this suppression may cause a decrease in cross-linkages within the cell wall architecture. The reduction in intra-network structures may contribute to keeping the cell wall loose under microgravity conditions.     

Hypergravity Provokes a Temporary Reduction in CD4+CD8+ Thymocyte Number and a Persistent Decrease in Medullary Thymic Epithelial Cell Frequency in Mice

Gravity change affects many immunological systems. We investigated the effects of hypergravity (2G) on murine thymic cells. Exposure of mice to 2G for three days reduced the frequency of CD4⁺CD8⁺ thymocytes (DP) and mature medullary thymic epithelial cells (mTECs), accompanied by an increment of keratin-5 and keratin-8 double-positive (K5⁺K8⁺) TECs that reportedly contain TEC progenitors. Whereas the reduction of DP was recovered by a 14-day exposure to 2G, the reduction of mature mTECs and the increment of K5⁺K8⁺ TEC persisted. Interestingly, a surgical lesion of the inner ear’s vestibular apparatus inhibited these hypergravity effects. Quantitative PCR analysis revealed that the gene expression of Aire and RANK that are critical for mTEC function and development were up-regulated by the 3-day exposure and subsequently down-regulated by the 14-day exposure to 2G. Unexpectedly, this dynamic change in mTEC gene expression was independent of the vestibular apparatus. Overall, data suggest that 2G causes a temporary reduction of DP and a persistent reduction of mature mTECs in a vestibular system-dependent manner, and also dysregulates mTEC gene expression without involving the vestibular system. These data might provide insight on the impact of gravity change on thymic functions during spaceflight and living.     

Deformation of the Outer Hair Cells and the Accumulation of Caveolin-2 in Connexin 26 Deficient Mice

BackgroundMutations in GJB2, which encodes connexin 26 (Cx26), a cochlear gap junction protein, represent a major cause of pre-lingual, non-syndromic deafness. The degeneration of the organ of Corti observed in Cx26 mutant—associated deafness is thought to be a secondary pathology of hearing loss. Here we focused on abnormal development of the organ of Corti followed by degeneration including outer hair cell (OHC) loss.MethodsWe investigated the crucial factors involved in late-onset degeneration and loss of OHC by ultrastructural observation, immunohistochemistry and protein analysis in our Cx26-deficient mice (Cx26^f/fP0Cre).ResultsIn ultrastructural observations of Cx26^f/fP0Cre mice, OHCs changed shape irregularly, and several folds or notches were observed in the plasma membrane. Furthermore, the mutant OHCs had a flat surface compared with the characteristic wavy surface structure of OHCs of normal mice. Protein analysis revealed an increased protein level of caveolin-2 (CAV2) in Cx26^f/fP0Cre mouse cochlea. In immunohistochemistry, a remarkable accumulation of CAV2 was observed in Cx26^f/fP0Cre mice. In particular, this accumulation of CAV2 was mainly observed around OHCs, and furthermore this accumulation was observed around the shrunken site of OHCs with an abnormal hourglass-like shape.ConclusionsThe deformation of OHCs and the accumulation of CAV2 in the organ of Corti may play a crucial role in the progression of, or secondary OHC loss in, GJB2-associated deafness. Investigation of these molecular pathways, including those involving CAV2, may contribute to the elucidation of a new pathogenic mechanism of GJB2-associated deafness and identify effective targets for new therapies.     

Yeast Cell Wall Extract Induces Disease Resistance against Bacterial and Fungal Pathogens in Arabidopsis thaliana and Brassica Crop

Housaku Monogatari (HM) is a plant activator prepared from a yeast cell wall extract. We examined the efficacy of HM application and observed that HM treatment increased the resistance of Arabidopsis thaliana and Brassica rapa leaves to bacterial and fungal infections. HM reduced the severity of bacterial leaf spot and anthracnose on A. thaliana and Brassica crop leaves with protective effects. In addition, gene expression analysis of A. thaliana plants after treatment with HM indicated increased expression of several plant defense-related genes. HM treatment appears to induce early activation of jasmonate/ethylene and late activation of salicylic acid (SA) pathways. Analysis using signaling mutants revealed that HM required SA accumulation and SA signaling to facilitate resistance to the bacterial pathogen Pseudomonas syringae pv. maculicola and the fungal pathogen Colletotrichum higginsianum. In addition, HM-induced resistance conferred chitin-independent disease resistance to bacterial pathogens in A. thaliana. These results suggest that HM contains multiple microbe-associated molecular patterns that activate defense responses in plants. These findings suggest that the application of HM is a useful tool that may facilitate new disease control methods.     

Selection of Orthologous Genes for Construction of a Highly Resolved Phylogenetic Tree and Clarification of the Phylogeny of Trichosporonales Species

The order Trichosporonales (Tremellomycotina, Basidiomycota) includes various species that have clinical, agricultural and biotechnological value. Thus, understanding why and how evolutionary diversification occurred within this order is extremely important. This study clarified the phylogenetic relationships among Tricosporonales species. To select genes suitable for phylogenetic analysis, we determined the draft genomes of 17 Trichosporonales species and extracted 30 protein-coding DNA sequences (CDSs) from genomic data. The CDS regions of Trichosporon asahii and T. faecale were identified by referring to mRNA sequence data since the intron positions of the respective genes differed from those of Cryptococcus neoformans (outgroup) and are not conserved within this order. A multiple alignment of the respective gene was first constructed using the CDSs of T. asahii, T. faecale and C. neoformans, and those of other species were added and aligned based on codons. The phylogenetic trees were constructed based on each gene and a concatenated alignment. Resolution of the maximum-likelihood trees estimated from the concatenated dataset based on both nucleotide (72,531) and amino acid (24,173) sequences were greater than in previous reports. In addition, we found that several genes, such as phosphatidylinositol 3-kinase TOR1 and glutamate synthase (NADH), had good resolution in this group (even when used alone). Our study proposes a set of genes suitable for constructing a phylogenetic tree with high resolution to examine evolutionary diversification in Trichosporonales. These can also be used for epidemiological and biogeographical studies, and may also serve as the basis for a comprehensive reclassification of pleomorphic fungi.     

Function and Evolutionary Origin of Unicellular Camera-Type Eye Structure

The ocelloid is an extraordinary eyespot organelle found only in the dinoflagellate family Warnowiaceae. It contains retina- and lens-like structures called the retinal body and the hyalosome. The ocelloid has been an evolutionary enigma because of its remarkable resemblance to the multicellular camera-type eye. To determine if the ocelloid is functionally photoreceptive, we investigated the warnowiid dinoflagellate Erythropsidinium. Here, we show that the morphology of the retinal body changed depending on different illumination conditions and the hyalosome manifests the refractile nature. Identifying a rhodopsin gene fragment in Erythropsidinium ESTs that is expressed in the retinal body by in situ hybridization, we also show that ocelloids are actually light sensitive photoreceptors. The rhodopsin gene identified is most closely related to bacterial rhodopsins. Taken together, we suggest that the ocelloid is an intracellular camera-type eye, which might be originated from endosymbiotic origin.