Sialyl Lewisx expression at the invasive front as a predictive marker of liver recurrence in stage II colorectal cancer.

Sialyl Lewisx (SLX) is a carbohydrate ligand for endothelial selectin that participates in cell adhesion, proliferation and scattering. It plays an important role in cancer cell adhesion to vascular endothelial cells, leading to hematogenous metastasis. The prognostic significance of SLX expression level at the invasive front in patients with stage II colorectal cancer (CRC) was examined. A total of 209 patients with stage II CRC curatively resected between 1997 and 2000 were enrolled. The preoperative serum SLX levels measured by radioimmunoassay and SLX immunoexpression levels at the invasive front, and at the non-invasive frontal region determined by tissue microarray were analyzed. SLX expression at the invasive front was positively associated with tumor invasion depth (P=0.007) and tumor budding grade (P=0.038). Disease-free survival curves differed between the high and low SLX-expression groups (5-year survival rates, 77.0 and 89.7%, respectively; P=0.036). Liver cancer recurrence was more frequent in the high-expression group than in the low-expression group (15.9 and 2.4%; P=0.002). Multivariate analysis revealed that its expression (hazard ratio, 5.26; P=0.015) and venous invasion (hazard ratio, 4.14; P=0.040) were independent predictive markers of liver cancer recurrence. Neither the preoperative serum SLX level nor SLX expression at the non-invasive frontal region showed any association with histopathological features or disease-free survival. SLX expression level at the invasive front is a promising marker for identifying patients with stage II CRC with a high risk of liver cancer recurrence.

Differential clinical benefits of 5-fluorouracil-based adjuvant chemotherapy for patients with stage III colorectal cancer according to CD133 expression status.

OBJECTIVE: CD133 has been recently identified as a marker of putative cancer stem cells in colorectal tumors. The ability of cancer stem cells to resist chemotherapy was clinically highlighted; however, whether CD133 expression can predict chemoresistance remains controversial. The objective of the study was to determine the relationship between clinical benefits of adjuvant chemotherapy and CD133 expression status in colorectal cancer. METHODS: We enrolled 234 patients with Stage III colorectal cancer who underwent curative resection. Among them, 149 received 5-fluorouracil-based adjuvant chemotherapy (chemotherapy group) and 85 did not (surgery-alone group). We immunohistochemically stained the specimens for CD133 on specimens evaluated the benefits of adjuvant chemotherapy according to CD133 expression using the Kaplan-Meier method and log-rank test. RESULTS: A comparison of disease-free survival between both the groups revealed a significant 3-year disease-free survival benefit of adjuvant chemotherapy in CD133-negative (92.2% versus 74.5%; P = 0.004), but not in CD133-positive patients (46.8% versus 52.9%; P = 0.67). Multivariate analysis corroborated the benefits of adjuvant chemotherapy in CD133-negative (P = 0.003, hazard ratio = 0.26), but not in CD133-positive patients. CONCLUSIONS: CD133-positive patients showed resistance to 5-FU-based chemotherapy, while CD133-negative patients experienced significant survival benefits from adjuvant chemotherapy not shared by CD133-positive patients.

Impact of race/ethnicity on prognosis in patients who underwent surgery for colon cancer: analysis for white, African, and East Asian Americans.

PURPOSE: We retrospectively investigated the impact of race/ethnicity on prognosis in patients who underwent surgery for colon cancer. METHODS: Surveillance, Epidemiology, and End Results population-based data on 39,210 colon cancer patients without distant metastasis who underwent radical surgery were analyzed. Prognostic impact of race/ethnicity for non-Hispanic white, Hispanic white, African American, and East Asian (Japanese, Chinese, Korean) American patients, and confounding factors of age, sex, registry region, year of diagnosis, tumor, node, metastasis system stage, tumor grade, tumor site, and the number of lymph nodes examined were analyzed by the Cox proportional hazard model. The lymph node count was analyzed and adjusted means were calculated by a generalized multiple regression model with respect to race and other factors. RESULTS: Significant differences due to race/ethnicity were observed in crude hazard ratios with respect to overall and colon cancer-specific mortality, which persisted even after adjusting for confounding factors. Adjusted hazard ratios of colon cancer-specific mortality for non-Hispanic white, Hispanic white, African American, and East Asian American patients were 1 (reference), 1.01 (95% confidence interval 0.91-1.12), 1.40 (95% confidence interval 1.31-1.50), and 0.83 (95% confidence interval 0.74-0.94), respectively. There were significant differences in crude number of lymph nodes examined among races, which were no longer significant after adjusting for covariates. CONCLUSIONS: East Asian American patients had significantly better prognosis, while African American patients had worse prognosis than non-Hispanic white patients, despite the identical adjusted number of lymph nodes examined after surgery for colon cancer. This disparity in prognosis among races/ethnicities should be taken into consideration when deciding adjuvant chemotherapy for nonwhite patients.

Histological comparison of long-bone cortex between 11-year-old giant cow with dermal dysplasia and the child cow aged 8.5 years.

Young calves are known to be formed with laminar bone in long-bone cortex during growing periods and the osteon formation begins later. Previously, we reported that an 11-year-old giant Holstein cow with dermal dysplasia showed a delayed osteon formation. An 8.5-year-old cow, born from the giant Holstein cow, also showed some dermal dysplasia and the outer-half layer of the child almost retained laminar bone similar to that of the mother, although the body weight was approximately normal. The mother had formed the inner circumferential lamella and the child was going to form the inner circumferential lamella, but their outer circumferential lamellas were not formed yet in both of them, when compared with a 12-years-old cow as a control of the mother. Therefore, we suggest on long-bone formation pattern that the child resembled the mother rather than the control, and that the child had more or less succeeded to the mother genes of delayed osteon formation as well as dermal dysplasia which seemed to be genetic collagen disorder, although there were mild gene appearances.

[A case of long-term survival after resection of advanced undifferentiated rectal cancer treated by short-term preoperative chemoradiotherapy and postoperative adjuvant chemotherapy].

A 75-year-old man with advanced undifferentiated rectal cancer, diagnosed by endoscopic biopsy, underwent preoperative short-term chemoradiotherapy (whole pelvis, 4 Gy × 5 day with UFT 400 mg/day × 7 day). Tumor size and lymph node swellings were reduced after radiation therapy. Down-staging was achieved from cT3, cN2, cStage III b to cT3, cN1, cStage III a. A curative low anterior resection with D3 lymphadenectomy including lateral lymph node dissection, was performed 4 weeks after the completion of chemoradiotherapy. Pathological findings of resected specimen showed undifferentiated carcinoma with regional lymph node involvement (pT2, pN1, pStage III a). The histological change in response to chemoradiation was evaluated as Grade 2. The postoperative course was uneventful and postoperative adjuvant chemotherapy (UFT+Uzel) was performed for six months (5 courses). No sign of recurrence has been found until 51 months after the operation. Undifferentiated rectal cancer is a rare condition with extremely poor prognosis according to the Japanese literature. Nine cases have been reported so far with only one long-term survivor. This combination of preoperative short-term chemoradiotherapy and adjuvant chemotherapy, which is one of the standard strategies for advanced rectal cancer in Western countries, but not common in Japan, may be a promising option for treatment of undifferentiated rectal cancer.

Application of a reservoir-type calcitriol transdermal patch in dairy cattle.

Our objective was to evaluate the effects of a reservoir-type calcitriol (5 mg/animal) transdermal patch on plasma calcitriol and calcium (Ca) concentrations in dairy cattle. A group of six heifers received three different types of patches: an encapsulated reservoir solution containing calcitriol alone (CAL), calcitriol with dodecylamine (CAL+D), or vehicle (CONT). The patches were applied to the skin of the tail for duration of 2 days at intervals of at least 3 weeks. Cattle that received CAL or CAL+D showed significant increases in plasma calcitriol and Ca concentrations on day 2 and days 2 and 3, respectively. The areas under the plasma concentration-time curve (AUC) values for plasma calcitriol and Ca in the CAL and CAL+D groups increased significantly compared to the CONT group. The data of this study describe evidence of measurable transdermal absorption of exogenous calcitriol from a patch and its sufficient biological action to elevate plasma Ca concentrations.

The evaluation of the curd forming ability of milk replacers.

Inadequate milk curd formation in the abomasum of newborn calves causes malnutrition and diarrhea. In order to define the factors of inadequate curd formation, we compared the curd forming ability among 9 kinds of milk replacers, bulk milk (raw milk), and skim milk both in vitro and in vivo. When rennet was added, the raw milk and one milk replacer formed firm curds. The rest of the milk replacers and skim milk did not form any curd. When a solution of HCl was added, raw milk, three milk replacers and skim milk formed the curd at pH 4.5, but the other milk replacers did not. When HCl was added following the rennet, raw milk, one milk replacer and skim milk formed the curd. In vivo, raw milk, two milk replacers and skim milk showed good curd formation whereas the other milk replacers showed poor curd formation inside the abomasums of the calves. This study showed that most of the milk replacers sold in Japan could not form the curd with rennet.

Molecular screening of canine GM1 gangliosidosis using blood smear specimens after prolonged storage: detection of carriers among shiba dogs in northern Japan.

Molecular screening of GM1 gangliosidosis in Shiba dogs was carried out in northern Japan using blood smear specimens after prolonged storage. Of 125 specimens obtained from 3 veterinary teaching hospitals for this screening, 68 specimens (54%) were adequate for direct amplification in a polymerase chain reaction (PCR)-based DNA test, and the percentage of adequacy was different at each hospital (34%, 73%, and 100%), suggesting that the amount of blood on the smear and the storage condition of specimens may affect adequacy. Of the 68 dogs examined, 2 dogs (2.9%) were heterozygous carriers for this disease and the other dogs were all genotypically normal. The results suggest blood smear specimens can be useful for PCR testing after prolonged storage provided specimens contain a generous amount of blood and have been adequately stored. The study also suggests that GM1 gangliosidosis may be widely prevalent in the Shiba dog population in northern Japan.

The interaction between orally administered non-steroidal anti-inflammatory drugs and prednisolone in healthy dogs.

The interaction between oral non-steroidal anti-inflammatory drugs (NSAIDs) and prednisolone administered concurrently for 30 days was studied in 18 healthy dogs divided into 3 groups of 6 dogs each: a drug-free negative control group (NC group) given 2 gelatin capsules; a group given meloxicam (0.1 mg/kg) and prednisolone (0.5 mg/kg) (MP group); and a group given a reduced dosage of ketoprofen (0.25 mg/kg, p.o.) and prednisolone (0.5 mg/kg, p.o.) (KP group). The dogs were periodically monitored by physical examinations, blood analyses, endoscopic examinations, fecal occult blood tests, renal function tests [effective renal plasma flow (ERPF) and glomerular filtration rate (GFR)], urinalyses [urinary sediments, and urinary micro-albumin to creatinine ratio (UAlb/Cre)], urinary enzyme indices, and haemostatic function tests [buccal mucosa bleeding time (BMBT), cuticle bleeding time (CBT)]. Significant changes were observed in the KP group, including a decrease of ERPF and GFR, an increased UAlb/Cre ratio, prolonged BMBT and CBT, as well as the presence of more severe grades of endoscopic lesions and fecal occult blood. In both the MP and KP groups, abnormal enzymuria with exfoliation of renal tubular epithelial cells in the urine was found. However, no significant changes in any of the other tests were observed in the MP group compared with the NC group. These findings suggest that the combination of NSAIDs, even selective COX-2 inhibitors, with prednisolone may be contraindicated due to the potential for serious adverse effects on the kidneys, the platelets, and the gastrointestinal tract.

Calcium metabolism of pregnant rats fed a vitamin D-depleted diet.

The effects of vitamin D (VD) deficiency on calcium (Ca) metabolism during pregnancy were evaluated in rats fed VD-repleted (VD-repleted rats) and VD-depleted (VD-depleted rats) diets. In the VD-depleted rats, the plasma concentrations of 1,25-dihydroxyvitamin D and Ca decreased severely, whereas the parathyroid hormone concentrations increased. The Ca contents of the feces of the VD-depleted rats were higher than those of the VD-repleted rats. The fetal Ca contents of the VD-repleted and VD-depleted rats increased continuously, but that of the VD-depleted rats was lower. These data reveal that VD deficiency during pregnancy induces severe hypocalcemia due to reduced intestinal absorption of Ca and elevated fetal demand for Ca.

Biomarkers for the activation of calcium metabolism in dairy cows: elevation of tartrate-resistant acid phosphatase activity by lowering dietary cation-anion difference is associated with the prevention of milk fever.

In our previous study, it was demonstrated that the administration of anion salts, which slightly lower the dietary cation-anion difference (DCAD), in the prepartum period is safe and effective for preventing milk fever in multiparous cows. In the present study, several biomarkers, which might show activation of Ca metabolism, were analyzed using stored samples in the previous study to investigate the mechanism of the preventive effect on milk fever by lowering DCAD. Changes in bone-specific alkaline phosphatase activity, osteocalcin and insulin-like growth factor I concentrations in serum were almost the same among the three groups of multiparous cows with or without the oral administration of anion salts, while the levels of these serum biomarkers in the group of primiparous cows (heifer group) were much higher compared with those in the three multiparous groups throughout the experimental period. Urinary deoxypyridinoline excretion was not a useful biomarker for dairy cows because it hardly changed during the peripartum period in all groups. However, serum tartrate-resistant acid phosphatase (TRAP) activity, which is known as a biomarker of osteoclast activity, was well associated with the administration of anion salts lowering DCAD because among the three multiparous groups, only the group of multiparous cows fed the anion salts (anion group) showed an increased level, which rose to the level in the heifer group, and was markedly higher than those in the other control groups of multiparous cows. The increased activity of serum TRAP in the anion group suggested that Ca in the plasma pool was mobilized smoothly from bone-bound Ca via mature osteoclasts at parturition, which might be due to prior activation under mild acidosis induced by slightly lowering DCAD. Therefore, TRAP was the best biomarker to monitor the activation of Ca metabolism in dairy cows fed anion salts.

Canine serum thermostable alkaline phosphatase isoenzyme from a dog with hepatocellular carcinoma.

A dog histopathologically diagnosed with hepatocellular carcinoma (HCC) showed very high serum alkaline phosphatase (ALP) activity. A supernatant of ascitic fluid and tumor tissue extracted from the dog also showed much higher ALP activity than normal. ALP isoenzyme analysis of samples was performed using polyacrylamide gel disk electrophoresis, and a wide, broad abnormal band was observed. By various treatments, the abnormal band showed thermostability, which is a characteristic of tumor-associated ALP that has only been reported in humans. The thermostable ALP isoenzyme was not found in sera from 39 dogs with several types of tumor that originated from the liver, except for HCC, nor was it found in 10 dogs with hepatic diseases that did not include hepatic tumors. The thermostable ALP isoenzyme seemed to be associated with canine HCC.

Changes of serum alkaline phosphatase activity in dry and lactational cows.

Serum alkaline phosphatase (ALP) activities were measured during dry and lactational periods to investigate the influence of lactation on serum ALP activity in cows. Higher levels of serum ALP activity were seen in lactational periods than in dry periods. The serum activities of bone-specific ALP (BALP), liver ALP (LALP), tartrate resistant acid phosphatase (TRAP) and aspartate aminotransferase also increased in lactational periods. ALP activities in the bone extract and in whey were decreased at similar rates by the addition of lectin. Moreover, since the ALP band in whey was observed to have the same migration in polyacrylamide gel (PAG) disk electrophoresis as that of the bone extract, analysis of ALP isoenzymes by lectin affinity or PAG disk electrophoresis could not distinguish ALP originating from the mammary gland from that of bone. In this study, it was clear that the increased level of serum ALP activity was due to increases of BALP and LALP in lactational periods. However, the extent of the influence of ALP originating from the mammary glands on serum ALP activity was unknown. Judging from changes of BALP and TRAP activities in the serum and the correlation between the both, it was guessed that ALP originating from the mammary glands influenced serum ALP activity.

Calcium metabolism in cows receiving an intramuscular injection of 1,25-dihydroxyvitamin D3 combined with prostaglandin F(2alpha) closely before parturition.

To determine the effect of exogenous 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] combined with induced parturition on calcium (Ca) metabolism, cows received a single intramuscular injection of 1,25(OH)2D3 and prostaglandin F(2alpha) (PGF(2alpha)) closely before calving. Ten late-pregnant, multiparous Holstein cows were assigned to 1,25(OH)2D3 group (five treated with both 1,25(OH)2D3 and PGF(2alpha)) and control group (five treated with PGF(2alpha)). 1,25(OH)2D3 group showed an increase in plasma Ca concentration around parturition, whereas control group revealed a decrease in plasma Ca level. Plasma Ca concentration in 1,25(OH)2D3 group were significantly higher than that in control group during -0.5 to 3 days after parturition.

Reduction of concanavalin A-induced expression of interferon-gamma by bovine lactoferrin in feline peripheral blood mononuclear cells.

Lactoferrin (LF), a glycoprotein present in milk, mucosal secretions and neutrophils, contributes to host defense and immunomodulation. In the present study, we investigated the effect of bovine LF (bLF) on cytokine messenger RNA (mRNA) expression in concanavalin A (ConA)-stimulated feline peripheral blood mononuclear cells (PBMC). Semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR) and real-time PCR showed a ConA-induced increase of interferon-gamma (IFN-gamma) mRNA expression but not of interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha) and IL-12 p40 mRNA in feline PBMC. This ConA-induced increase of IFN-gamma mRNA expression was inhibited by addition of bLF not only 30 min before ConA stimulation but also 10, 20 and 40 min after ConA stimulation. Western blotting showed that protein tyrosine kinase (PTK) and extracellular signal-regulated kinase (ERK) in feline PBMC were activated within 10 min after the ConA stimulation and that the activation of both kinases had almost disappeared by 40 min after stimulation. Moreover, the ConA-induced IFN-gamma mRNA expression was partly prevented by genistein, a global PTK inhibitor, and PD-98059, an ERK inhibitor, respectively. These results suggest that bLF is able to inhibit the ConA-induced IFN-gamma mRNA expression by abrogation of intracellular signaling activated after interaction between ConA and its receptor.

Preliminary study of histological comparison on the growth patterns of long-bone cortex in young calf, pig, and sheep.

Some young large farm animals show a laminar bone formation in the long-bone cortex. Such a laminar bone is gradually replaced by Haversian bone with osteons during their growth periods. In this preliminary study, we observed the transverse ground samples of tibia cortex in young calves, pigs, and sheep by backscattered electron imaging. The cortex bones of all the newborn (NB) animals were basically formed with laminar bone structures. The NB and 1-month-old (1-M) calves had a typical concentric structure of laminar bone, whereas the NB and 1-M pigs showed a wire-netting bone with laminar-bone units. The NB sheep was similar to the calf rather than the pig. In the growth rate of bone volume, sheep was similar to calf up to 6 months after birth (6-M). Such calf and sheep showed a more rapid ratio of bone volume than pig. A few osteons had initially appeared in the innermost layer of the 6-M calf. A 1-year-old (1-Y) calf showed scattered osteons in the bone cortex, but many laminar-bone units were still retained in the outer layer. A 6-M pig had many osteons in the entire cortex but only a few osteons in the outermost layer. In the 6-M sheep, no osteons were observed, whereas a 1-Y sheep showed a relatively small number of osteons mainly in the middle layer but a higher osteon-volume than the 1-Y calf. In the 1-Y sheep, the more widely absorbed areas by bone-remodeling with osteons were observed as compared with the 1-Y calf, and the bone volume was decreased from the 6-M into the 1-Y sheep because of the remarkable bone-absorption. Thus, calf kept on possessing many laminar-bone units for a longer time in the growth period than sheep, while pig showed the earliest bone-remodeling with osteons. These results may be caused by their different body size and withers height in calf and sheep after growing and the difference of the dependence upon mother's body during juvenile period between pig and calf with sheep. The initial region of osteon formation may be distinguishable among their animals, respectively. However, further detailed investigations of their young animals at successive stages will be necessary.

The plasma superoxide scavenging activity in canine cancer and hepatic disease.

To clarify the relationship between plasma antioxidant activity and diseases in dogs, plasma samples were collected from 6 healthy dogs and 16 diseased dogs (6 dogs with cancer, 5 dogs with hepatic disease, and 5 dogs with inflammation ), and measured superoxide anion scavenging activities. Antioxidant activities of canine plasma were evaluated by measuring their superoxide anion (O(2)(-.)) scavenging activities with electron spin response spectroscopy combined with spin trapping reagent, 5,5-dimethyl-1-pyrroline-N-oxide (DMPO). Total O(2)(-.) scavenging activities in the presence of plasma of diseased dogs tended to be higher than those in healthy controls, especially significant higher activities in the presence of canine plasma of hepatic disease and inflammation were observed. In diseased dogs, KCN-insensitive activities, suggesting the activity of manganese-containing superoxide dismutase (Mn-SOD), were significantly higher than those in healthy controls. Therefore, it seems that there is a possibility of utilizing of plasma O(2)(-.) scavenging activity as one of clinical indicators for oxidative-related diseases such as cancer, hepatic disease and inflammation in dogs.

Association between changes in plasma calcium concentration and plasma tartrate-resistant acid phosphatase activity in periparturient cows.

Eight periparturient Holstein Friesian cows were examined for plasma tartrate-resistant acid phosphatase (TRAP) activity to assess the degree of bone metabolic activity and to evaluate the association between the change in calcium (Ca) concentration and bone metabolism during the periparturient period. Milk fever occurred in 1 of 8 cows just after parturition. Plasma TRAP activities did not markedly change in 5 of 8 cows during the experimental period. The changing rate of Ca between preparturition and just after parturition was under -20% in 3 of 8 cows, and low TRAP activities were observed in 2 of these 3 cows. This study suggests that cows with a low TRAP activity are at risk of developing milk fever in comparison to cows with high TRAP activity. Temporary increases of parathyroid hormone were observed in 7 cows, but not in the cow with milk fever.

Serum activities of tartrate-resistant acid phosphatase and bone specific alkaline phosphatase as indices of bone metabolism in the cow.

The correlation between the serum hydroxyproline concentration and serum activity levels of TRAP and BALP was examined in 41 cows. The correlated coefficient (r) was 0.6391 for TRAP and 0.3147 for BALP, respectively. Judging from the significant correlation to the serum hydroxyproline concentration, serum TRAP activity is an index for bone metabolism in cows. Serum TRAP activity was therefore measured in 205 healthy cows (2-9 years old) in order to observe the changes in bone resorption with aging and milk production. TRAP levels differed slightly between group A (< or =4 yrs) and B (5 yrs< or =) at the same stage of lactation. The activity levels rose slightly at the height of lactation stage and during the dry stage.

Genomic characterization of TT viruses (TTVs) in pigs, cats and dogs and their relatedness with species-specific TTVs in primates and tupaias.

Using PCR with primers derived from a non-coding region of the human TT virus (TTV) genome, the TTV sequence in serum samples obtained from pigs (Sus domesticus), dogs (Canis familiaris) and cats (Felis catus) was identified and the entire genomic sequence was determined for each representative isolate. Three TTV isolates (Sd-TTV31 from a pig, Cf-TTV10 from a dog and Fc-TTV4 from a cat) comprising 2878, 2797 and 2064 nucleotides, respectively, each had three open reading frames (ORFs) encoding 436-635 (ORF1), 73-105 (ORF2) and 224-243 (ORF3) aa but lacked ORF4, similar to tupaia TTV. ORF3 was presumed to arise from a splicing of TTV mRNA, similar to human prototype TTV. Although the nucleotide sequence of Sd-TTV31, Cf-TTV10 and Fc-TTV4 differed by more than 50% from each other and from previously reported TTVs of 3.4-3.9 kb and TTV-like mini viruses (TLMVs) of 2.8-3.0 kb isolated from humans and non-human primates as well as tupaia TTVs of 2.2 kb, they resembled known TTVs and TLMVs with regard to genomic organization and presumed transcriptional profile rather than animal circoviruses of 1.7-2.3 kb. Phylogenetic analysis revealed that Sd-TTV31, Cf-TTV10 and Fc-TTV4 were closer to TTVs from lower-order primates and tupaias than to TTVs from higher-order primates and TLMVs. These results indicate that domestic pigs, cats and dogs are naturally infected with species-specific TTVs with small genomic size and suggest a wide distribution of TTVs with extremely divergent genomic sequence and length in animals.