Analysis of sequential ruminal temperature sensor data from dairy cows to identify cow subgroups by clustering and predict calving through supervised machine learning.

The present study investigated the applicability of a calving prediction model based on supervised machine learning of ruminal temperature (RT) data in dairy cows. The existence of cow subgroups for prepartum RT changes was also examined, and the predictive performance of the model was compared among these subgroups. RT data were collected from 24 Holstein cows at 10 min intervals using an RT sensor system. The average hourly RT was calculated and data were expressed as residual RTs (rRT = actual RT - mean RT for the same time on the previous three days). The mean rRT decreased beginning at approximately 48 h before calving to a low of -0.5°C at 5 h before calving. However, two cow subgroups were identified: cows with a late and small rRT decrease (Cluster 1, n = 9) and those with an early and large rRT decrease (Cluster 2, n = 15). A calving prediction model was developed using five features extracted from the sensor data (indicative of prepartum rRT changes) through a support vector machine. Cross-validation showed that calving within 24 h was predicted with a sensitivity of 87.5% (21/24) and precision of 77.8% (21/27). A significant difference in sensitivity was observed between Clusters 1 and 2 (66.7 vs. 100%, respectively), while none was observed for precision. Therefore, the model based on RT data with supervised machine learning has the potential to efficiently predict calving, although improvements for specific cow subgroups are required.

Deciphering two rounds of cell lineage segregations during bovine preimplantation development.

Blastocyst formation gives rise to the inner cell mass (ICM) and trophectoderm (TE) and is followed by the differentiation of the epiblast (Epi) and primitive endoderm (PrE) within the ICM. Although these two-round cell lineage differentiations underpin proper embryogenesis in every mammal, their spatiotemporal dynamics are quite diverse among species. Here, molecular details of the blastocyst stage in cattle were dissected using an optimized in vitro culture method. Blastocyst embryos were placed on agarose gel filled with nutrient-rich media to expose embryos to both gaseous and liquid phases. Embryos derived from this "on-gel" culture were transferred to surrogate mothers on day (D) 10 after fertilization and successfully implanted. Immunofluorescent studies using on-gel-cultured embryos revealed that the proportion of TE cells expressing the pluripotent ICM marker, OCT4, which was beyond 80% on D8, was rapidly reduced after D9 and reached 0% on D9.5. This first lineage segregation process was temporally parallel with the second one, identified by the spatial separation of Epi cells expressing SOX2 and PrE cells expressing SOX17. RNA-seq comparison of TE cells from D8 in vitro fertilized embryos and D14 in vivo embryos revealed that besides drastic reduction of pluripotency-related genes, TE cells highly expressed Wnt, FGF, and VEGF signaling pathways-related genes to facilitate the functional maturation required for feto-maternal interaction. Quantitative PCR analysis of TE cells derived from on-gel culture further confirmed time-dependent increments in the expression of key TE markers. Altogether, the present study provides platforms to understand species-specific strategies for mammalian preimplantation development.

Relationship between the timing of insemination based on estrus detected by the automatic activity monitoring system and conception rates using sex-sorted semen in Holstein dairy cattle.

We investigated the optimal timing of artificial insemination (AI) for achieving pregnancy according to the onset/end of estrus detected by an accelerometer system in Holstein cattle. The conception rates of conventional semen were used as a reference. The conception rate from AI of sex-sorted semen was higher at -4 to 4 h (57.1%) from the end of estrus than those at -12 to -4 h (37.7%) and 12-20 h (30.3%), whereas AI at 4-12 h showed an intermediate conception rate (47.4%). Conversely, conception rates were similar in AI performed between 0 and 32 h from the onset of estrus. Regarding conventional semen, the interval from the onset and end of estrus did not affect conception rates. The present results suggest that the time of the end of estrus is the better indicator of optimal AI timing for sex-sorted semen than the onset of estrus.

Generation of viable calves derived from developmentally mature blastocysts produced by on-gel culture.

Conventional culture systems for bovine embryos are unable to support sustained embryonic development until the developmentally mature blastocyst stage. Although we have previously developed an on-gel culture system that enables bovine blastocysts to complete cell segregation events at day (D) 10 following in vitro culture, the development of D10 blastocysts to term has yet to be achieved. In this study, we attained full-term development of D10 mature blastocysts produced using an on-gel culture system. Two calves derived from on-gel-cultured embryos were vaginally born, showing normal birth and placental weights and no obvious morphological abnormalities. Moreover, we detected no abnormalities in blood metabolic profile analyses. Our findings indicate that on-gel culturing can be used to facilitate the development of developmentally mature blastocysts to term, and produce healthy viable calves. This culture system could make a valuable contribution to cattle production and would enable a range of analyses for characterizing bovine-specific pre-implantation development.

Higher susceptibility to osmolality of the medaka (Oryzias latipes) mutants in orthologue genes of mammalian skin transglutaminases.

Transglutaminase (TG) is an essential enzyme to catalyze cross-linking reactions of epidermal proteins. Recently, we biochemically characterized human skin TG orthologues for medaka (Oryzias latipes), a model fish. By genome editing, gene-modified fishes for the two orthologues were obtained, both of which lack the ordinal enzymes. These fish appeared to exhibit higher susceptibility to osmolality at the period of larvae.

The effectiveness of dapagliflozin for sleep-disordered breathing among Japanese patients with obesity and type 2 diabetes mellitus.

Weight reduction is important in patients with sleep-disordered breathing (SDB). In Japanese patients, slight weight reduction is effective for improving the severity of SDB. However, the effect of weight reduction after administration of sodium glucose co-transporter 2 (SGLT2) inhibitor for SDB remains unclear. The aim of this study was to evaluate the improvement of SDB from baseline after administration of dapagliflozin (5 mg) once daily for 24 weeks among Japanese patients with obesity and type 2 diabetes mellitus. Thirty Japanese patients with type 2 diabetes mellitus and SDB were enrolled in a 24-week, prospective, open-label, single-arm, multicentre trial. SDB was defined as at least five 3% oxygen desaturation index (ODI) events per hour, and moderate to severe SDB was defined as at least 15 ODI events per hour. The primary endpoint was the change in 3% ODI between before dapagliflozin administration and at 24 weeks. The prevalence of moderate to severe SDB was 20% in the present study. After administration of dapagliflozin, fasting glucose, HbA1c, aspartate aminotransferase, total cholesterol, low-density lipoprotein cholesterol, and estimated globular filtration rate decreased significantly. The improvement of 3% ODI was observed in patients with moderate to severe SDB but not mild SDB (from 25.0 ± 3.8 at baseline to 18.5 ± 6.1 at 24 weeks, p = 0.017). In conclusion, dapagliflozin might improve moderate to severe SDB but not mild SDB in Japanese patients with obesity and type 2 diabetes mellitus.

[Case of ischemic ileitis complicated by portal venous gas].

A 65-year-old woman was admitted with sudden-onset abdominal pain. Abdominal computed tomography revealed hepatic portal venous gas. Physical and laboratory examination suggested that a conservative approach was appropriate; however, 4 days later, the pain recurred and severe ischemic enteritis was diagnosed. A stenosis was identified 60 cm distal to the start of the ileum, and partial resection of the small intestine was performed. The diagnosis of ischemic enteritis was confirmed. Ischemic enteritis affecting the small intestine is uncommon, and enteritis causing intestinal stenosis with hepatic portal vein gas is even rarer.

Effect of heat exposure on the growth and developmental competence of bovine oocytes derived from early antral follicles.

In dairy cows, low fertility caused by summer heat stress continues into the cooler autumn season. This can be caused by impaired oocyte quality in small growing follicles during summer. Here, we subjected oocyte-cumulus-granulosa complexes (OCGCs) derived from early antral follicles (0.5-1 mm) to in vitro growth (IVG) culture under two different temperature settings (the control and heat shock groups), and evaluated effects of heat exposure on growth and developmental competence of oocytes, factors affecting the developmental competence of oocytes (steroidogenesis of granulosa cells, oxidative stress in oocytes, and cell-to-cell communication between oocytes and somatic cells). Oocyte diameters after culture were smaller in the heat shock group. Although nuclear maturation and cleavage rates were similar between the groups, blastocyst rates were lower in the heat shock group (0.0%) than in the control group (27.7%), and reduced glutathione (GSH) levels in oocytes were lower in the heat shock group. Supplementation of cysteine, which stimulates GSH synthesis, increased GSH level and improved blastocyst rate of heat shocked oocytes (27.9%). These results suggest that heat exposure impairs the growth and developmental competence of oocytes in early antral follicles through GSH depletion, which can induce low fertility during summer and the following autumn.

Low oxygen environment and astaxanthin supplementation promote the developmental competence of bovine oocytes derived from early antral follicles during 8 days of in vitro growth in a gas-permeable culture device.

We evaluated the effects of a constant low (5-5%) and modulated (5-20%) oxygen environments on the in vitro development of bovine oocyte-cumulus-granulosa cell complexes (OCGCs) cultured in the presence or absence of an antioxidant (astaxanthin: Ax). OCGCs were cultured in a gas permeable culture device for 8 days in 5-5% O2 (±Ax) and 5-20% O2 (±Ax) culture conditions. In the oxygen modulated culture conditions, the oxygen concentration was switched from 5% to 20% on day 4 of culture. Ax promoted the viability of OCGCs (P < 0.05), but both oxygen and Ax had a significant effect on ROS production levels by OCGCs (P < 0.05). Specifically, ROS levels were significantly lower and higher under 5-5% O2 (+Ax) and 5-20% O2 (-Ax) conditions, respectively (P < 0.05), with intermediate levels observed in the 5-5% O2 (-Ax) and the 5-20% O2 (+Ax) culture conditions. The steroidogenic pattern was characterized by increasing estradiol-17ß but with constant progesterone production levels regardless of culture conditions, suggesting the inhibition of luteinization-like changes in granulosa cells. OCGCs cultured in the 5-20% O2 (+Ax) had higher nuclear maturation rates (P < 0.05) that were similar to the oocytes grown in vivo. However, there was no clear difference in the subsequent cleavage rates among the 5-5% O2 (±Ax) and the 5-20% O2 (+Ax) culture conditions (P > 0.05). A constant low oxygen environment significantly promoted the blastocyst rates (P < 0.05); however, the presence of Ax in the 5-20% O2 (+Ax) condition also promoted development similar to the OCGCs cultured in the 5-5% O2 (-Ax) condition (P > 0.05). In conclusion, exposure of OCGCs to constant low oxygen or oxygen modulation in the presence of Ax promotes the healthy development of OCGCs during the 8-day IVG culture using the gas permeable culture device.

Simultaneous free fatty acid elevations and accelerated desaturation in plasma and oocytes in early postpartum dairy cows under intensive feeding management.

A severe negative energy balance and high circulating free fatty acids (FFA) in postpartum cows impair fertility. The lipotoxicity of FFA has been shown to decrease the quality of bovine oocytes in vitro. Therefore, excess FFA in cells is converted to triacylglycerol (TAG), a non-toxic form, to avoid lipotoxicity. We recently reported that the TAG content in oocytes was higher in postpartum lactating cows subjected to grazing management than in heifers (Theriogenology 176: 174-182, 2021). The present study investigated the compositions of the energy metabolism-related lipids, FFA and TAG, in the plasma and oocytes of cows at different lactation stages under indoor intensive feeding management in order to obtain insights into lipotoxicity in oocytes, particularly those in early postpartum cows. Blood and oocytes were collected from 20 lactating cows categorized into the following lactation groups: 20-30 days in milk (DIM) (n = 5), 40-50 DIM (n = 5), 60-80 DIM (n = 5), and 130-160 DIM (n = 5). Daily energy balance data were obtained for 3 weeks prior to oocyte collection using the ovum pick up (OPU) method. The contents and compositions of FFA and TAG in plasma and oocytes were analyzed using liquid chromatography-mass spectrometry. As expected, plasma FFA was high at 20-30 DIM, decreased by 50 DIM, and was maintained at a low level for the remainder of the experimental period. Similar changes were observed in oocyte FFA and TAG with DIM as plasma FFA. Oocyte FFA positively correlated with plasma FFA (P < 0.05), but negatively correlated with the mean energy balance 1 and 21 days before OPU (P < 0.05). Relationships were noted between the composition and content of FFA in plasma and oocytes, with the FFA 16:1/16:0 and 18:1/18:0 ratios positively correlating with the total amount of FFA (P < 0.05). Elevated oocyte FFA in cows in the early postpartum period under intensive feeding management suggested that oocytes were at a high risk of FFA lipotoxicity. Furthermore, the present results implied that the severe negative energy balance in the previous few weeks was closely related to increases in oocyte FFA, which supports the importance of long-term cow feeding management for preserving the quality of oocytes in the early postpartum period. The present results provide insights into the effects of high circulating FFA on the fertility of postpartum cows.

Postpartum cows showed high oocyte triacylglycerols concurrently with high plasma free fatty acids.

Impaired oocyte quality is one of the main causes of low fertility in modern high-yielding dairy cows. One of the potential factors of the impaired oocyte quality is the effects of free fatty acids (FFA). In fact, high FFA supplementation to culture media exacerbated oocyte developmental competence in vitro. Meanwhile, artificially induced high blood FFA levels in heifers did not affect the lipid composition of oocytes in vivo; however, the oocyte lipid profile of postpartum cows has not yet been investigated. Therefore, the profile of lipids involved in energy metabolism, including FFA and triacylglycerols (TAG), and their relationship between plasma and oocytes were compared among cows at different lactation stages. Heifers were used as a control group that was not affected by lactation. Plasma and oocytes were collected from heifers (n = 4) and 14 Holstein cows categorized to the early lactation stage: 25-47 days in milk (DIM) (n = 6), peak lactation stage: 61-65 DIM (n = 4), and middle lactation stage: 160-202 DIM (n = 4). The FFA and TAG profiles of plasma and oocytes were examined by liquid chromatography mass spectrometry. Plasma FFA positively correlated with oocyte TAG (P < 0.05). Plasma FFA and oocyte TAG were significantly higher in cows in the early lactation stage than in heifers (P < 0.05), while the peak and middle lactation stage groups had intermediate levels. The proportion of oleic acid in plasma increased concurrently with elevations in total FFA, while the compositions of oocyte FFA and TAG fatty acyls were constant regardless of plasma FFA concentration or oocyte TAG content. The present results suggest that high postpartum plasma FFA concentrations affect the quantity of oocyte TAG. Taken together with the adverse effects of high FFA concentrations on oocyte developmental competence in vitro, oocyte quality in postpartum cows may be impaired due to high circulating FFA concentrations. These results provide a more detailed understanding of the effects of postpartum high circulating FFA concentrations on the low fertility of cows.

Relationship between the timing of the first postpartum ovulation and antral follicle counts in Holstein cows.

BACKGROUND: The timing of the first postpartum ovulation is an important factor affecting the timing of estrous resumption in dairy cows. The first postpartum ovulation is delayed in cows producing large amounts of milk with an intensive negative energy balance. The antral follicle count (AFC) and serum anti-Müllerian hormone concentrations are known to be indicators of the ovarian reserve, which is the number and quality of follicles left in a pair of ovaries and known as an indicator of female fertility. Cows with higher AFC have been proven to show higher pregnancy rate and shorter calving to conception intervals; however, the relationship between the timing of the first postpartum ovulation and ovarian reserve remains unclear. Therefore, this study examined the relationships between postpartum follicular dynamics, the ovarian cycle, nutritional status, and ovarian reserve. METHODS: Transrectal ultrasonography was conducted from calving to 70-120 days in milk (DIM) in 26 cows to monitor AFC, follicular dynamics and the ovarian cycle. Body weight (BW) and milk yield were used as indicators of nutritional status. RESULTS: The first postpartum ovulation was significantly later in cows with low AFC (< 25) than in those with high AFC (≥25), while changes in BW from calving to the nadir and milk production were similar in both groups. The present results also suggested that cows with low AFC and a delayed first postpartum ovulation had a shorter first ovarian cycle after the first postpartum ovulation. The mean DIM of the first postpartum artificial insemination (AI) and days open (days from calving to AI with which pregnancy was achieved) were similar in high and low AFC groups. CONCLUSIONS: The first postpartum ovulation was significantly earlier in cows with high AFC than in those with low AFC. The assumed reason for this result was higher sensitivity to luteinizing hormone and larger androstenedione and estradiol production in follicles in high AFC cows. Therefore, cows with high AFC may be more fertile than those with low AFC while their milk production increase and BW decrease; it means they are in negative energy balance. (340/350 words).

Effect of summer heat stress on gene expression in bovine uterine endometrial tissues.

Heat stress negatively affects reproductive functions in cows. Increased temperature disturbs fetal development in utero. However, the effect of heat stress on uterine endometrial tissues has not been fully examined. Using qPCR analysis, we measured the mRNA expression of various molecular markers in uterine endometrial tissue of dairy cows from Hokkaido, Japan, in winter and summer. Markers examined were heat shock proteins (HSPs), antioxidant enzymes (catalase, copper/zinc superoxide dismutase, manganese superoxide dismutase, and glutathione peroxidase 4), inflammatory cytokines, and interferon stimulated genes. Our results showed heat stress, body and milk temperatures were higher during summer than during winter. Expression levels of HSP27, HSP60, and HSP90 mRNA, and of catalase and copper/zinc superoxide dismutase mRNA were lower in summer than in winter. Tumor necrosis factor alpha expression was higher in summer than in winter. In conclusion, summer heat stress may reduce the expression of HSPs, affecting the levels of inflammatory cytokines in bovine uterine endometrial tissue.

Lipidomic profiling of dairy cattle oocytes by high performance liquid chromatography-high resolution tandem mass spectrometry for developmental competence markers.

A comparative lipidomic profiling analysis of dairy cattle oocytes with different developmental competences was performed using a combination of high performance liquid chromatography-high resolution tandem mass spectrometry and multivariate statistical analysis. Significant lipidomic changes were identified in degenerating oocytes. Total triacylglycerol in the degenerating oocytes was 1.8-fold higher than that in the normal oocytes; however, total cardiolipin was 53.5% lesser than that in the normal oocytes, which indicated attenuation of energy metabolism. Compared to those in the normal oocytes, triacylglycerols in the degenerating oocytes were composed of longer and more unsaturated acyl chains. In contrast, the acyl chains in free fatty acids present in the degenerating oocytes were shorter and with lesser degree of unsaturation compared to those in the normal oocytes. Moreover, a significant decrease in degenerating oocytes were found in total phosphatidylinositol (14.8 ± 7.6 pmol vs. 24.8 ± 5.5 pmol), total phosphatidylcholine (20.8 ± 8.7 pmol vs. 33.5 ± 7.2 pmol), and total plasmalogen ethanolamine (9.0 ± 4.7 pmol vs. 16.8 ± 5.2 pmol), which indicated dysfunction of lipid-metabolizing enzymes in oocytes during degeneration. Thus, increase of triacylglycerols together with the decrease of certain phospholipid species could be potential markers of oocyte developmental competence. In addition to providing a new approach to investigate the lipidomic changes in oocyte development, the lipidomic profiling in the present study has revealed insights that hold potential to unravel the role of lipid metabolism in oocyte developmental competence in cattle.

[Clinical analysis of primary lung cancer with a thin-walled cavity to explain the mechanism of thin-walled cavity formation].

We report 8 rare cases of primary lung cancer which showed a thin-walled cavity on chest X-ray and CT. We analyzed 8 cases (7 men, 1 woman) of primary lung cancer with thin-walled cavities admitted to our hospital between 1995 and 2006. The subjects were aged between 45 and 84 years of age (median: 72 years old). The reason for detection was treatment for tuberculosis in 1 case, ileus owing to metastasis to the small intestine in 1 case and tension pnumathorax 1 case, while 5 cases had abnormal chest x-ray film shadows without symptoms. Histologically, there were 5 cases of adenocarcinoma, 2 of squamous cell carcinoma, and 1 of adenosquamous cell carcinoma. Though various reports on the mechanism of the development of thin-walled cavity formation have been made, we suggest that it mainly develops by a check-valve mechanism, based on evaluation of the clinical course.

Acute manganese administration alters dopamine transporter levels in the non-human primate striatum.

We used positron emission tomography (PET) to measure non-invasively the effect of acute systemic administration to manganese sulfate (MnSO4) on dopamine transporter (DAT) levels in the living non-human primate brain. Baboons received [11C]-WIN 35,428 PET scans to measure DAT levels before and after acute MnSO4 administration. In one animal, we observed a 46% increase in DAT binding potential (BP), a measure of DAT binding site availability, 1 week after Mn administration. DAT levels returned to baseline values at 4 months and remained constant at 10 months after treatment. A subsequent single MnSO4 injection to the same animal also resulted in a 57% increase in DAT-BP, 2 days after administration. In a second animal, a 76% increase in DAT-BP relative to baseline was observed at 3 days after Mn injection. In this animal, the DAT-BP returned to baseline levels after 1 month. Using in vitro receptor binding assays, we found that Mn inhibits [3H]-WIN 35,428 binding to rat striatal DAT with an inhibitory constant (Ki) of 2.0+/-0.3mM (n=4). Saturation isotherms and Scatchard analysis of [3H]-WIN 35,428 binding to rat striatal DAT showed a significant decrease (30%, p<0.001) in the maximal number of binding sites (Bmax) in the presence of 2mM MnSO4. No significant effect of Mn was found on binding affinity (Kd). We also found that Mn inhibits [3H]-dopamine uptake with an IC50 of 11.4+/-1.5mM (n=4). Kinetic studies and Lineweaver-Burk analysis showed a significant decrease (40%, p<0.001) in the maximal velocity of uptake (Vmax) with 5mM MnSO4. No significant effect of Mn was found on Michaelis-Menten constant (Km). These in vitro findings suggest that the increase in DAT levels in vivo following acute Mn administration may be a compensatory response to its inhibitory action on DAT. These findings provide helpful insights on potential mechanisms of Mn-induced neurotoxicity and indicate that the DAT in the striatum is a target for Mn in the brain.

Clinical characteristics of autoimmune hepatitis in older aged patients.

BACKGROUND AND AIMS: Autoimmune hepatitis (AIH) is an inflammatory disease of the liver that usually develops in middle-aged women. However, due to the increasing aging of the population and better diagnostic facilities, AIH is now diagnosed in older patients as well. This analysis compared the clinical and pathologic characteristics of older and middle-aged patients with AIH. PATIENTS AND METHODS: Thirteen older patients with AIH (mean age, 75.0+/-5.3 years; range, 70-89 years) and 27 middle-aged patients (mean age, 51.3+/-5.8 years; range, 41-60 years) were included in this study. In addition, the use of different treatment regimens, including prednisolone therapy and ursodeoxycholic acid (UDCA), was examined. RESULTS: There were no significant differences in gender, complications of other autoimmune diseases, and liver function tests between groups. However, the degree of hepatic fibrosis was significantly higher in older patients compared with middle-aged patients (P<0.05). Four patients with AIH in the older age group were successfully managed by UDCA alone. CONCLUSION: This study shows that older patients with abnormal liver function should be checked for AIH. In addition, UDCA may be an effective drug for management of older patients with AIH.

Neuronal differentiation is associated with a redox-regulated increase of copper flow to the secretory pathway.

Brain development requires a fine-tuned copper homoeostasis. Copper deficiency or excess results in severe neuro-pathologies. We demonstrate that upon neuronal differentiation, cellular demand for copper increases, especially within the secretory pathway. Copper flow to this compartment is facilitated through transcriptional and metabolic regulation. Quantitative real-time imaging revealed a gradual change in the oxidation state of cytosolic glutathione upon neuronal differentiation. Transition from a broad range of redox states to a uniformly reducing cytosol facilitates reduction of the copper chaperone Atox1, liberating its metal-binding site. Concomitantly, expression of Atox1 and its partner, a copper transporter ATP7A, is upregulated. These events produce a higher flux of copper through the secretory pathway that balances copper in the cytosol and increases supply of the cofactor to copper-dependent enzymes, expression of which is elevated in differentiated neurons. Direct link between glutathione oxidation and copper compartmentalization allows for rapid metabolic adjustments essential for normal neuronal function.

Effect of stent implantation on upstream coronary artery compliance--a cause of late plaque rupture?

Stent implantation in the rabbit aorta has been shown to increase vessel wall compliance at the inflow to the stent, but it is uncertain whether similar effects might be seen in the coronary arteries of humans and whether this would have any significant clinical consequences. First, we measured vessel compliance (systolic lumen area--diastolic lumen area/pulse pressure) before, immediately after, and at the 6-month follow-up visit at a site 5 mm upstream of the proximal edge of an implanted coronary stent in patients undergoing coronary intervention using motorized pull-back intravascular ultrasound recordings. Compliance in the upstream segment increased significantly immediately after stenting (before 7.13 +/- 1.49 vs after 10.73 +/- 1.36 mm2/mm Hg, p = 0.03), an effect that was unchanged at 6 months of follow-up (11.84 +/- 2.11 mm2/mm Hg, p = 0.08 vs before stenting). Second, we examined the site of plaque rupture in all patients presenting with an acute coronary syndrome in whom the culprit lesion was in a vessel that had had a stent implanted >12 months previously (n = 31). Plaque rupture was statistically more likely at the inflow to the stent (n = 22) than at other sites within the culprit vessel (n = 9, p <0.01). We conclude that stenting causes an increase in vessel compliance immediately proximal to the stent, and that when a vessel has been previously stented, plaque rupture is most likely to occur at the stent inflow site.

Biochemical Characterization of Medaka (Oryzias latipes) Transglutaminases, OlTGK1 and OlTGK2, as Orthologues of Human Keratinocyte-Type Transglutaminase.

Calcium-dependent transglutaminases (TGs) are a family of enzymes that catalyze protein cross-linking and/or attachment of primary amines in a variety of organisms. Mammalian TGs are implicated in multiple biological events such as skin formation, blood coagulation, and extracellular matrix stabilization. Medaka (Oryzias latipes) has been used as a model fish to investigate the physiological functions of mammalian proteins. By analysis of the medaka genome, we found seven TGs orthologues, some of which apparently corresponded to the mammalian TG isozymes, TG1, TG2, and Factor XIII. All orthologues had preserved amino acid residues essential for enzymatic activity in their deduced primary structures. In this study, we analyzed biochemical properties of two orthologues (OlTGK1 and OlTGK2) of mammalian epithelium-specific TG (TG1) that are significantly expressed at the transcriptional level. Using purified recombinant proteins for OlTGK1 and OlTGK2, we characterized their catalytic reactions. Furthermore, immunohistochemical analyses of fish sections revealed higher expression in the pancreas (OTGK1), intervertebral disk (OlTGK2) and pharyngeal teeth (OlTGK2) as well as in the skin epidermis.