Porphyromonas gingivalis infection alters microRNA composition in extracellular vesicles.

OBJECTIVES: Periodontitis, commonly associated with Porphyromonas gingivalis (Pg), involves intricate alterations of oral intercellular interactions, in which extracellular vesicles (EVs) play a pivotal role. The understanding of the miRNA profiles in the EVs derived from Pg-infected cells (Pg-EVs) remains incomplete despite acknowledging their importance in intercellular communication during periodontitis. Therefore, our objective was to identify and characterize the miRNAs enriched in Pg-EVs. METHODS: Microarray analysis was conducted to examine the miRNA profiles in the EVs derived from Pg-infected THP-1 cells. We compared the identified miRNAs with those upregulated in the EVs after stimulation with LPS. Additionally, we explored how inhibiting TLR signaling during Pg infection affects the transcription of specific miRNAs. We investigated the unique sequence motifs specific to the miRNAs concentrated in Pg-EVs. RESULTS: The levels of eleven miRNAs, including miR-155, were increased in Pg-EVs compared with those elevated after LPS stimulation. The Pg-induced miR-155 upregulation via TLR2 but not TLR4 signaling suggests the influence of TLR signaling on the miRNA composition of EVs. Furthermore, the miRNAs upregulated in Pg-EVs contained AGAGGG and GRGGSGC sequence motifs. CONCLUSIONS: Our findings demonstrate that Pg-induced alterations in EV-containing miRNA composition occur in a TLR4-independent manner. Notably, the concentrated miRNAs in Pg-EVs harbor specific motifs with a high G + C content within their sequences. The upregulation of specific miRNAs in EVs under infectious conditions suggests the influence of both innate immune receptor signals and miRNA sequence characteristics.

Transcriptome Analysis by RNA Sequencing of Mouse Embryonic Stem Cells Stocked on International Space Station for 1584 Days in Frozen State after Culture on the Ground.

As a space project, in "Stem Cells" by the Japan Aerospace Exploration Agency (JAXA), frozen mouse ES cells were stored on the International Space Station (ISS) in the Minus Eighty Degree Laboratory Freezer for ISS (MELFI) for 1584 days. After taking these cells back to the ground, the cells were thawed and cultured, and their gene expressions were comprehensively analyzed using RNA sequencing in order to elucidate the early response of the cells to long-time exposure to space radiation consisting of various ionized particles. The comparisons of gene expression involved in double-stranded break (DSB) repair were examined. The expressions of most of the genes that were involved in homologous recombination (HR) and non-homologous end joining (NHEJ) were not significantly changed between the ISS-stocked cells and ground-stocked control cells. However, the transcription of Trp53inp1 (tumor protein 53 induced nuclear protein-1), Cdkn1a (p21), and Mdm2 genes increased in ISS-stocked cells as well as Fe ion-irradiated cells compared to control cells. This suggests that accumulated DNA damage caused by space radiation exposure would activate these genes, which are involved in cell cycle arrest for repair and apoptosis in a p53-dependent or -independent manner, in order to prevent cells with damaged genomes from proliferating and forming tumors.

Incidence of Radiation-induced Nausea and Vomiting: A Prospective Single-institution Pilot Study.

Radiation-induced nausea and vomiting (RINV) is a frequent adverse event that occurs in patients undergoing radiotherapy. However, research on RINV is underrepresented. This prospective single-institution exploratory pilot study investigated the incidence of RINV according to the irradiation site and observed the efficacy of symptomatic antiemetic treatment in controlling symptoms of RINV. The primary outcomes were the proportions of emesis-free days and nausea-free days. The secondary endpoints included the time to the first episode of RINV, frequency of vomiting, and severity of nausea, including its impact on eating habits and weight loss. Fifteen patients were enrolled in each group (minimal, low, and moderate emetogenic risk). All patients received greater than 20 Gy in five fractions. Evaluation was based on weekly questionnaires completed by patients during routine clinic visits. Nausea and vomiting occurred in 11 and 0 patients, respectively. Six of 15 patients in the minimal-risk group, 1 in the low-risk group, and 4 in the moderate-risk group experienced nausea. Although all 11 symptomatic patients were offered antiemetics, only 3 used them, who reported satisfactory control of nausea. The percentage of emesis-free days for all patients was 100% and the percentage of nausea-free days for the 11 patients who developed RINV was 38%. An unexpectedly high percentage of patients in the minimal-risk group experienced nausea; all had breast cancer. Future studies should investigate factors beyond the irradiation site, including the characteristics of the patient and the treatment, to better predict an individual's risk of RINV.

Factors associated with quality of life in patients receiving palliative radiotherapy for bone metastases: a secondary cross-sectional analysis of data from a prospective multicenter observational study.

OBJECTIVE: To identify factors significantly associated with quality of life (QOL) and determine if these associations are strong enough to predict certain aspects of QOL without measuring them. METHODS: We conducted an exploratory secondary analysis of baseline data of 224 patients (enrolled between December 2020 and March 2021) from a previously published prospective observational study on radiotherapy for bone metastases at 26 centres. Using univariable linear regression, we assessed the association between patient/treatment factors and QOL scale scores as measured by the European Organization for Research and Treatment of Cancer (EORTC) QOL Questionnaire Core 15-Palliative (QLQ-C15-PAL) and the EORTC QOL Questionnaire Bone Metastases module (QLQ-BM22). RESULTS: Age and sex were not significantly associated with QOL. Worse performance status, higher pain scores, and opioid and single-fraction use were significantly associated with most QOL scales; these four factors were associated with worse global QOL, worse functioning status, and more severe symptoms. The coefficients of determination for most QOL scales were less than 0.2, indicating that most of the variability in QOL scores was not explained by any of the explanatory variables. CONCLUSION: Performance status, pain intensity, and opioid and single-fraction use were significantly associated with most QOL scales. However, the associations were not strong enough to estimate QOL. ADVANCES IN KNOWLEDGE: To date, the association between treatment factors and QOL in patients with bone metastases has not been fully studied. We identified the factors that were significantly associated with QOL and found that these associations were not strong enough to predict QOL.

Evaluation of dose perturbations around iodine-125 seed sources in supplemental external beam prostate radiotherapy.

We investigated dose perturbations caused by 125I seeds in patients undergoing supplemental external beam radiotherapy (EBRT) for prostate cancer. We examined two types of nonradioactive seed models: model 6711 and model STM1251. All experiments were performed using a water-equivalent phantom. Radiochromic film was used to measure the dose distributions adjacent to the seeds upstream and downstream of the external beam source. Single and clusters of multiple seeds were placed in slots in a solid water (SW) slab to measure dose perturbations with separate versus dense seed placement at beam energies of 6 or 10 MV. Monte Carlo simulations (MCSs) were also performed to include the theoretical basis against film dosimetry. Distinct patterns of dose enhancement (buildup [BU]) were upstream, and dose reduction (builddown [BD]) were downstream of the radiation source. Model 6711 with lower photon beam energies produced larger dose perturbations of BU and BD than the model STM1251. The results showed the same tendency with different seed placements and beam energies. However, these differences were not observed in the rotational irradiation measurement, which replicated a clinical plan. Dose perturbations around seeds result in dose enhancement and dose reduction with varying impact depending on the photon beam energy and seed type. This has the potential to cancel out these perturbations using multiple beam direction fields.

Income and Employment of Patients at the Start of and During Follow-up After Palliative Radiation Therapy for Bone Metastasis.

Purpose: The aim of this study was to understand the income and employment status of patients at the start of and during follow-up after palliative radiation therapy for bone metastasis. Methods and Materials: From December 2020 to March 2021, a prospective multi-institutional observational study was conducted to investigate income and employment of patients at the start of administration of radiation therapy for bone metastasis and at 2 and 6 months after treatment. Of 333 patients referred to radiation therapy for bone metastasis, 101 were not registered, mainly because of their poor general condition, and another 8 were excluded from the follow-up analysis owing to ineligibility. Results: In 224 patients analyzed, 108 had retired for reasons unrelated to cancer, 43 had retired for reasons related to cancer, 31 were taking leave, and 2 had lost their jobs at the time of registration. The number of patients who were in the working group was 40 (30 with no change in income and 10 with decreased income) at registration, 35 at 2 months, and 24 at 6 months. Younger patients (P = 0), patients with better performance status (P = 0), patients who were ambulatory (P = .008), and patients with lower scores on a numerical rating scale of pain (P = 0) were significantly more likely to be in the working group at registration. There were 9 patients who experienced improvements in their working status or income at least once in the follow-up after radiation therapy. Conclusions: The majority of patients with bone metastasis were not working at the start of or after radiation therapy, but the number of patients who were working was not negligible. Radiation oncologists should be aware of the working status of patients and provide appropriate support for each patient. The benefit of radiation therapy to support patients continuing their work and returning to work should be investigated further in prospective studies.

Porphyromonas gingivalis outer membrane vesicles in cerebral ventricles activate microglia in mice.

OBJECTIVE: Porphyromonas gingivalis (Pg) is thought to be involved in the progression of Alzheimer's disease (AD). Whether Pg or its contents can reach the brain and directly affect neuropathology is, however, unknown. Here, we investigated whether outer membrane vesicles (OMVs) of Pg translocate to the brain and induce the pathogenic features of AD. MATERIAL AND METHODS: Pg OMVs were injected into the abdominal cavity of mice for 12 weeks. Pg OMV translocation to the brain was detected by immunohistochemistry using an anti-gingipain antibody. Tau protein and microglial activation in the mouse brain were examined by western blotting and immunohistochemistry. The effect of gingipains on inflammation was assessed by real-time polymerase chain reaction using human microglial HMC3 cells. RESULTS: Gingipains were detected in the region around cerebral ventricles, choroid plexus, and ventricular ependymal cells in Pg OMV-administered mice. Tau and phosphorylated Tau protein increased and microglia were activated. Pg OMVs also increased the gene expression of proinflammatory cytokines in HMC3 cells in a gingipain-dependent manner. CONCLUSION: Pg OMVs, including gingipains, can reach the cerebral ventricle and induce neuroinflammation by activating microglia. Pg OMVs may provide a better understanding of the implications of periodontal diseases in neurodegenerative conditions such as AD.

Comparison of biological measurement and physical estimates of space radiation in the International Space Station.

Nowadays, ordinary people can travel in space, and the possibility of extended durations in an environment such as moon of the Earth and Mars with higher space radiation exposures compared to past missions, is increasing. Until now, the physical doses of space radiation have been measured, but measurement of direct biological effects has been hampered by its low dose and low dose-rate effect. To assess the biological effects of space radiation, we launched and kept frozen mouse embryonic stem (ES) cells in minus eighty degree Celsius freezer in ISS (MELFI) on the International Space Station (ISS) for a maximum of 1,584 days. The passive dosimeter for life science experiments in space (PADLES) was attached on the surface of the sample case of the ES cells. The physical dosimeter measured the absorbed dose in water. After return, the frozen cells were thawed and cultured and their chromosome aberrations were analyzed. Comparative experiments with proton and iron ion irradiation were performed at particle accelerators on Earth. The wild-type ES cells showed no differences in chromosomal aberrations between the ground control and ISS exposures. However, we detected an increase of chromosome aberrations in radio-sensitized histone H2AX heterozygous-deficient mouse ES cells and found that the rate of increase against the absorbed dose was 1.54-fold of proton irradiation at an accelerator. On the other hand, we estimated the quality factor of space radiation as 1.48 ± 0.2. using formulas of International Commission of Radiation Protection (ICRP) 60. The relative biological effectiveness (RBE) observed from our experiments (1.54-fold of proton) was almost equal (1.04-fold) to the physical estimation (1.48 ± 0.2). It should be important to clarify the relation between biological effect and physical estimates of space radiation. This comparative study paves a way to reveal the complex radiation environments to reduce the uncertainty for risk assessment of human stay in space.

Galectin-3 promotes the adipogenic differentiation of PDGFRα+ cells and ectopic fat formation in regenerating muscle.

Worldwide prevalence of obesity is associated with the increase of lifestyle-related diseases. The accumulation of intermuscular adipose tissue (IMAT) is considered a major problem whereby obesity leads to sarcopenia and metabolic disorders and thus is a promising target for treating these pathological conditions. However, whereas obesity-associated IMAT is suggested to originate from PDGFRα+ mesenchymal progenitors, the processes underlying this adipogenesis remain largely unexplored. Here, we comprehensively investigated intra- and extracellular changes associated with these processes using single-cell RNA sequencing and mass spectrometry. Our single-cell RNA sequencing analysis identified a small PDGFRα+ cell population in obese mice directed strongly toward adipogenesis. Proteomic analysis showed that the appearance of this cell population is accompanied by an increase in galectin-3 in interstitial environments, which was found to activate adipogenic PPARγ signals in PDGFRα+ cells. Moreover, IMAT formation during muscle regeneration was significantly suppressed in galectin-3 knockout mice. Our findings, together with these multi-omics datasets, could unravel microenvironmental networks during muscle regeneration highlighting possible therapeutic targets against IMAT formation in obesity.

Extracellular vesicles of P. gingivalis-infected macrophages induce lung injury.

Periodontal diseases are common inflammatory diseases that are induced by infection with periodontal bacteria such as Porphyromonas gingivalis (Pg). The association between periodontal diseases and many types of systemic diseases has been demonstrated; the term "periodontal medicine" is used to describe how periodontal infection/inflammation may impact extraoral health. However, the molecular mechanisms by which the factors produced in the oral cavity reach multiple distant organs and impact general health have not been elucidated. Extracellular vesicles (EVs) are nano-sized spherical structures secreted by various types of cells into the tissue microenvironment, and influence pathophysiological conditions by delivering their cargo. However, a detailed understanding of the effect of EVs on periodontal medicine is lacking. In this study, we investigated whether EVs derived from Pg-infected macrophages reach distant organs in mice and influence the pathophysiological status. EVs were isolated from human macrophages, THP-1 cells, infected with Pg. We observed that EVs from Pg-infected THP-1 cells (Pg-inf EVs) contained abundant core histone proteins such as histone H3 and translocated to the lungs, liver, and kidneys of mice. Pg-inf EVs also induced pulmonary injury, including edema, vascular congestion, inflammation, and collagen deposition causing alveoli destruction. The Pg-inf EVs or the recombinant histone H3 activated the NF-κB pathway, leading to increase in the levels of pro-inflammatory cytokines in human lung epithelial A549 cells. Our results suggest a possible mechanism by which EVs produced in periodontal diseases contribute to the progression of periodontal medicine.

Conversion from epithelial to partial-EMT phenotype by Fusobacterium nucleatum infection promotes invasion of oral cancer cells.

The ability of cancer cells to undergo partial-epithelial mesenchymal transition (p-EMT), rather than complete EMT, poses a higher metastatic risk. Although Fusobacterium nucleatum mainly inhabits in oral cavity, attention has been focused on the F. nucleatum involvement in colorectal cancer development. Here we examined the p-EMT regulation by F. nucleatum in oral squamous cell carcinoma (OSCC) cells. We cultured OSCC cells with epithelial, p-EMT or EMT phenotype with live or heat-inactivated F. nucleatum. Expression of the genes involved in epithelial differentiation, p-EMT and EMT were examined in OSCC cells after co-culture with F. nucleatum by qPCR. Cell growth and invasion of OSCC cells were also examined. Both live and heat-inactivated F. nucleatum upregulated the expression of p-EMT-related genes in OSCC cells with epithelial phenotype, but not with p-EMT or EMT phenotype. Moreover, F. nucleatum promoted invasion of OSCC cells with epithelial phenotype. Co-culture with other strains of bacteria other than Porphyromonas gingivalis did not alter p-EMT-related genes in OSCC cells with epithelial phenotype. F. nucleatum infection may convert epithelial to p-EMT phenotype via altering gene expression in OSCC. Oral hygiene managements against F. nucleatum infection may contribute to reduce the risk for an increase in metastatic ability of OSCC.

Upregulation of PTK7 and ß-catenin after vaginal mechanical dilatation: an examination of fibulin-5 knockout mice.

INTRODUCTION AND HYPOTHESIS: Pelvic organ prolapse (POP) in women is associated with deficiency of elastic fibers, and fibulin-5 is known to be a critical protein in the synthesis of elastin. The purpose of this study is to investigate the related pathway for the synthesis of elastin via fibulin-5 using fibulin-5 knockout mice. METHODS: Fibulin-5 knockout mice were generated using the CRISPR/Cas9 system, and vaginal dilatation was used to mimic vaginal delivery. We divided the mice into three groups: Fbln5+/+ mice immediately after dilatation (Fbln5+/+ day0), Fbln5+/+ mice 3 days after dilatation (Fbln5+/+ day3) and Fbln5-/- mice 3 days after dilatation (Fbln5-/- day3). Proteins related to elastogenesis in the vaginal wall were measured by liquid chromatography mass spectrometry (LC-MS/MS) analysis, and differences in the expression of these proteins between the Fbln5-/- mice and the Fbln5+/+ mice were analyzed using western blotting. RESULTS: In the LC-MS/MS analysis, protein tyrosine kinase 7 (PTK7) was not detected in the Fbln5-/- day3 group, although the expression increased by > 1.5 times between the Fbln5+/+ day0 and day3 groups. PTK7 and ß-catenin are known to act in the Wnt/ß-catenin pathway, and both were upregulated after dilatation in the Fbln5+/+ mice, though not in the Fbln5-/- mice. CONCLUSION: Our findings suggest that these proteins are involved in elastogenesis via fibulin-5, and the impairment of these proteins might be the underlying cause of POP manifestation.

Outer membrane vesicles of Porphyromonas gingivalis attenuate insulin sensitivity by delivering gingipains to the liver.

Outer membrane vesicles (OMVs) are nanosized particles derived from the outer membrane of gram-negative bacteria. Oral bacterium Porphyromonas gingivalis (Pg) is known to be a major pathogen of periodontitis that contributes to the progression of periodontal disease by releasing OMVs. The effect of Pg OMVs on systemic diseases is still unknown. To verify whether Pg OMVs affect the progress of diabetes mellitus, we analyzed the cargo proteins of vesicles and evaluated their effect on hepatic glucose metabolism. Here, we show that Pg OMVs were equipped with Pg-derived proteases gingipains and translocated to the liver in mice. In these mice, the hepatic glycogen synthesis in response to insulin was decreased, and thus high blood glucose levels were maintained. Pg OMVs also attenuated the insulin-induced Akt/glycogen synthase kinase-3 ß (GSK-3ß) signaling in a gingipain-dependent fashion in hepatic HepG2 cells. These results suggest that the delivery of gingipains mediated by Pg OMV elicits changes in glucose metabolisms in the liver and contributes to the progression of diabetes mellitus.

Enhanced homologous recombination by the modulation of targeting vector ends.

The field of genome editing was founded on the establishment of methods, such as the clustered regularly interspaced short palindromic repeat (CRISPR) and CRISPR-associated protein (CRISPR/Cas) system, used to target DNA double-strand breaks (DSBs). However, the efficiency of genome editing also largely depends on the endogenous cellular repair machinery. Here, we report that the specific modulation of targeting vectors to provide 3' overhangs at both ends increased the efficiency of homology-directed repair (HDR) in embryonic stem cells. We applied the modulated targeting vectors to produce homologous recombinant mice directly by pronuclear injection, but the frequency of HDR was low. Furthermore, we combined our method with the CRISPR/Cas9 system, resulting in a significant increase in HDR frequency. Thus, our HDR-based method, enhanced homologous recombination for genome targeting (eHOT), is a new and powerful method for genome engineering.

The Benefits of Docetaxel Plus Cisplatin and 5-Fluorouracil Induction Therapy in Conversion to Curative Treatment for Locally Advanced Esophageal Squamous Cell Carcinoma.

BACKGROUND: Definitive chemoradiotherapy (CRT), used for treatment of patients with an initial diagnosis of unresectable locally advanced esophageal cancer, has led to unsatisfactory long-term prognosis. Moreover, CRT can lead to esophageal fistula, perforation, and strictures. Therefore, strong induction chemotherapeutic treatments are necessary to reduce the tumor volume for subsequent radical esophagectomy. This study aimed to determine the oncological utility of docetaxel plus cisplatin and 5-fluorouracil (DCF) and the technical feasibility of subsequent esophagectomy for locally advanced esophageal cancer. METHODS: Eighty-seven patients with clinical borderline unresectable T3 and T4 esophageal squamous cell carcinoma without distant metastases were included in this study. There were 44 patients in primary DCF group and 43 patients in definitive CRT group, and perioperative and long-term oncological outcomes were compared between the two groups. RESULTS: Twenty-two patients (50%) achieved R0 resection in the DCF group. Albeit not significant, the rate of curative treatment was higher in the DCF group than the definitive CRT group (p = 0.099). The overall survival (OS) and progression-free survival (PFS) were better with DCF than with definitive CRT (median OS, 29 vs. 17 months, p = 0.206; median PFS, 10 vs. 6 months, p = 0.020). Specifically, the OS of patients with a Charlson score of less than 3 among the DCF-treated patients tended to be better than those among the definitive CRT-treated patients. CONCLUSION: DCF and subsequent esophagectomy achieved R0 resection in 50% of the patients and was associated with better long-term oncological outcomes in patients with initially unresectable esophageal cancer if their systemic status is acceptable.

Postoperative Pneumonia is Associated with Long-Term Oncologic Outcomes of Definitive Chemoradiotherapy Followed by Salvage Esophagectomy for Esophageal Cancer.

BACKGROUND OR PURPOSE: As we previously indicated, postoperative pneumonia has a negative impact on the overall survival after planned esophagectomy. However, the impact of postoperative pneumonia after salvage esophagectomy on long-term oncologic outcomes still remains unclear. This study aimed to indicate the association between postoperative pneumonia and long-term outcomes of definitive chemoradiotherapy followed by salvage esophagectomy. Furthermore, we determined a prediction model for overall survival (OS) and disease-free survival (DFS) using a survival classification and regression tree (CART). METHODS: Ninety-three patients who underwent CRT followed by esophagectomy for thoracic esophageal cancer were identified for this study. Forty-nine patients and 44 patients were included in the salvage and neoadjuvant groups, respectively. We investigated the association between postoperative pneumonia and long-term oncologic outcomes following salvage esophagectomy. RESULTS: Patients from the salvage group tended to have a lower OS compared to neoadjuvant group (median survival: salvage, 24 months vs neoadjuvant, 43 months, p = 0.117). Multivariate analyses revealed that postoperative pneumonia adversely affected both OS (p < 0.001) and DFS (p = 0.044) after salvage esophagectomy. We generated the prediction model for OS and DFS in the salvage group using survival CART. Postoperative pneumonia was the most important parameter for predicting the OS. DISCUSSION: The present study demonstrates the long-term outcomes and risk factors for mortality of salvage esophagectomy. To improve OS after salvage surgery, the development of a means of decreasing pulmonary complications is needed.

Technical feasibility and oncologic safety of diagnostic endoscopic resection for superficial esophageal cancer.

BACKGROUND AND AIMS: Active use of endoscopic resection (ER) for cM3-SM2 esophageal cancer may enable sufficient extent of esophageal resection and help determine the need for lymph node dissection based on histopathologic findings. However, ER preceding esophagectomy may have an adverse impact on outcomes. This study was designed to determine the technical feasibility and oncologic safety of diagnostic ER. METHODS: A single-institution retrospective cohort study was performed between July 2008 and June 2014. During this period, 135 consecutive patients with clinical T1a-M3N0M0, T1b-SM1N0M0, and T1b-SM2N0M0 primary esophageal cancer were referred to our division. Eight patients who underwent chemoradiotherapy as primary treatment were excluded because of inadequate pathologic findings. Based on oncologic and physical factors, we categorized the remaining 127 patients into 2 groups: primary esophagectomy (n = 54) and primary ER (n = 73). RESULTS: In all 127 patients, the 3-year overall survival (OS) and disease-free survival (DFS) rates were 95.7% and 87.6%, respectively. No adverse event requiring surgical intervention was observed after ER. Diagnostic ER had no negative impact on surgical outcomes, DFS, and OS after esophagectomy. Fourteen patients (19.2%) of those who received primary ER underwent curative resection, whereas 11 (20.4%) who had pT1a disease, no lymphovascular invasion, and no pathologic lymph node metastasis underwent primary esophagectomy. CONCLUSIONS: Diagnostic ER for cM3-SM2 esophageal cancer with or without subsequent esophagectomy was feasible and safe, not only from a surgical perspective but also an oncologic perspective. Approximately 20% of cM3-SM2N0M0 patients can potentially avoid undergoing additional treatment including esophagectomy using diagnostic ER.

Low-dose irradiation of mouse embryos increases Smad-p21 pathway activity and preserves pluripotency.

PURPOSE: To study the outcomes of mouse preimplantation embryos irradiated with low doses of X-rays (≤ 1 Gy) and investigate apoptosis and pluripotency of the irradiated embryos. METHODS: Mouse embryos at the 2-cell stage were collected for in vitro culture. After reaching the 8-cell stage, embryos were irradiated with various low doses of X-rays (0-1 Gy). Blastocysts with a normal appearance were transferred into a pseudopregnant uterus. The developmental rate to blastocysts and the survival rate following embryo transfer were examined. Expression levels of p21, Smad2, Foxo1, Cdx2, Oct4, and Nanog genes were measured by RT-PCR. Apoptotic cells in mouse blastocysts were examined immunofluorescently by staining for cleaved caspase-3. RESULTS: More than 90% of non-irradiated and low-dose X-ray-irradiated preimplantation embryos developed to morphologically normal blastocysts that could be implanted and survive in the uterus. However, embryos irradiated with X-rays had more apoptotic cells in a dose-dependent manner. Expression of p21, Smad2, and Foxo1 genes in X-ray-irradiated embryos was increased significantly, while expression of Cdx2, Oct4, and Nanog genes was maintained in comparison with non-irradiated embryos. CONCLUSIONS: Although irradiated embryos contained apoptotic cells, the low doses of irradiation did not disturb development of 8-cell stage embryos to blastocysts or their survival in utero. The underlying mechanisms might involve anti-apoptotic systems, including the Smad-p21 pathway, and preservation of pluripotency.

Mutation-induced loss of APP function causes GABAergic depletion in recessive familial Alzheimer's disease: analysis of Osaka mutation-knockin mice.

The E693Δ (Osaka) mutation in APP is linked to familial Alzheimer's disease. While this mutation accelerates amyloid ß (Aß) oligomerization, only patient homozygotes suffer from dementia, implying that this mutation is recessive and causes loss-of-function of amyloid precursor protein (APP). To investigate the recessive trait, we generated a new mouse model by knocking-in the Osaka mutation into endogenous mouse APP. The produced homozygous, heterozygous, and non-knockin littermates were compared for memory, neuropathology, and synaptic plasticity. Homozygotes showed memory impairment at 4 months, whereas heterozygotes did not, even at 8 months. Immunohistochemical and biochemical analyses revealed that only homozygotes displayed intraneuronal accumulation of Aß oligomers at 8 months, followed by abnormal tau phosphorylation, synapse loss, glial activation, and neuron loss. These pathologies were not observed at younger ages, suggesting that a certain mechanism other than Aß accumulation underlies the memory disturbance at 4 months. For the electrophysiology studies at 4 months, high-frequency stimulation evoked long-term potentiation in all mice in the presence of picrotoxin, but in the absence of picrotoxin, such potentiation was observed only in homozygotes, suggesting their GABAergic deficit. In support of this, the levels of GABA-related proteins and the number of dentate GABAergic interneurons were decreased in 4-month-old homozygotes. Since APP has been shown to play a role in dentate GABAergic synapse formation, the observed GABAergic depletion is likely associated with an impairment of the APP function presumably caused by the Osaka mutation. Oral administration of diazepam to homozygotes from 6 months improved memory at 8 months, and furthermore, prevented Aß oligomer accumulation, indicating that GABAergic deficiency is a cause of memory impairment and also a driving force of Aß accumulation. Our findings suggest that the Osaka mutation causes loss of APP function, leading to GABAergic depletion and memory disorder when wild-type APP is absent, providing a mechanism of the recessive heredity.

Single-energy metal artifact reduction in postimplant computed tomography for I-125 prostate brachytherapy: Impact on seed identification.

PURPOSE: To evaluate the effectiveness of the single-energy metal artifact reduction (SEMAR) technique for improving the accuracy of I-125 seed identification in postimplant computed tomography (CT) after prostate brachytherapy. METHODS AND MATERIALS: Postimplant CT images of 40 patients treated with I-125 prostate brachytherapy were acquired. For all patients, 2 data sets were reconstructed, 1 with SEMAR algorithms (SEMAR image), and the other without SEMAR algorithms (non-SEMAR image). Seed locations are automatically detected by the automatic seed finder tool, and their locations were compared between the SEMAR and non-SEMAR images. Dosimetric parameters using seed locations as detected were compared. RESULTS: The true-positive fraction of properly detected seeds on the SEMAR image as determined from a reference seed distribution defined by one investigator was significantly higher than the true-positive fraction on the non-SEMAR image (p = 0.011). The variabilities in D90 (p = 0.001), V100 (p = 0.007), and V150 (p = 0.007) were significantly reduced for seed location on the SEMAR image as compared with non-SEMAR image. CONCLUSIONS: Prostate postimplant CT with SEMAR improved the accuracy of seed localization and postimplant dosimetric parameters.