Supplementation with difructose anhydride III promotes passive calcium absorption in the small intestine immediately after calving in dairy cows.

The incidence of hypocalcemia increases in high-parity dairy cows because resorption of bone Ca is delayed in these animals, and they appear to have a reduced ability to absorb Ca from the intestine during the early postpartum period. Difructose anhydride (DFA) III has been shown to promote the absorption of intestinal Ca via a paracellular pathway. However, past studies have not reported this effect in peripartum dairy cows. Therefore, we investigated the effect of DFA III supplementation on Ca metabolism during the peripartum period to determine whether DFA III promotes intestinal Ca absorption via this route. Seventy-four multiparous Holstein cows were separated into DFA and control groups based on their parity and body weight. The feed of the DFA group was supplemented with 40g/d of DFA III from -14 to 6d relative to calving. The control group did not receive DFA III. At calving (0h relative to calving), serum Ca declined below 9mg/dL in both groups. However, serum Ca concentrations were greater in the DFA group than in the control group at 6, 12, 24, and 48h relative to calving, and the time required for serum Ca to recover to 9mg/dL during the postpartum period was shorter in the high-parity cows in the DFA group than in those in the control group. Parathyroid hormone concentrations increased immediately after calving in both groups and were greater in the control group than in the DFA group at 12 and 24h relative to calving. Serum 1,25-dihydroxyvitamin D concentrations increased at 0 and 12h relative to calving in both groups and were higher in the control group than in the DFA group at 72h relative to calving. Serum concentrations of the bone-resorption marker cross-linked N-telopeptide of type I collagen (NTX) were not different between the groups during peripartum period, and serum NTX in all cows was lower at 0, 6, 12, 24, 48, and 72h relative to calving than at -21, 4, and 5d relative to calving. Thus, DFA treatment induced faster recovery of serum Ca, although bone resorption was restrained. In conclusion, DFA III promotes intestinal passive Ca absorption via the paracellular pathway during the early postpartum period; this absorption is unaffected by aging.

Short communication: Difructose anhydride III promotes calcium absorption from the duodenum in cattle.

Difructose anhydride (DFA) III promotes the intestinal absorption of calcium via a paracellular pathway in rats. In dairy cows, DFA III reaches the duodenum without being degraded by ruminal bacteria and hence could be used to control hypocalcemia. The aims of the present study were to investigate the percentage of DFA III that appears in the duodenum of cows and to determine the effect of DFA III on calcium absorption from duodenal fluid. The first experiment was performed in 3 ruminally and duodenally cannulated dry Holstein cows in a 3 × 3 Latin square design. Each experimental period lasted 7 d. On the first day, the cows were ruminally fed one of the following treatments: 0 (DFA0), 50 (DFA50), or 100 (DFA100) g/d of DFA III, using cobalt-EDTA as a liquid phase marker. Difructose anhydride III was detected in duodenal fluid 1 h after feeding, and its concentration peaked 4 h after feeding, in a dose-dependent manner. The percentages of DFA III that appeared in the duodenum after the DFA50 and DFA100 treatments were 69.1 ± 7.0% and 67.9 ± 5.6%, respectively. The second experiment used the everted duodenal sacs of cattle (n = 7 in each group). Sacs were incubated in artificial mucosal fluid containing 1 mM DFA III or no DFA III (control) for 60 min with 100% O2 in a water bath at 37 °C. After incubation, the calcium concentration of the artificial serosal fluid in the everted sacs was measured. Calcium absorption was higher in the DFA III-treated group than in the control group (803 ± 161 and 456 ± 74 nmol/cm of sac, respectively). The above results demonstrate that approximately 70% of administered DFA III reached the duodenum of cows intact. Moreover, similar to its effects on calcium absorption in rats, DFA III promoted calcium absorption via a paracellular pathway in the duodenum of cows.

Changes of Serum Calcium Concentration, Frequency of Ruminal Contraction and Feed Intake Soon after Parturition of Dairy Cows Fed Difructose Anhydride III.

Requirements to control the large decrease in serum calcium (Ca) due to parturition and to increase the feed intake soon after parturition have been well accepted in dairy cows. This study was aimed to investigate the feed intake affected by serum Ca concentration with difructose anhydride (DFA) III supplement in dairy cows soon after parturition. Fourteen transition Holstein cows were divided into DFA and control (CONT) groups within 1 to 5 parity variations in each group. Measurement schedule for an individual cow was from 14 d before parturition to 7 d following parturition. The cows in DFA group were supplied 0.2 kg/head/d of DFA III feed containing 40 g of pure DFA III while the cows in CONT group received no DFA III. Other feeding procedures were the same for all cows in both groups. At parturition (d 0), serum Ca concentration sharply declined in both groups (p<0.05). Time interval for recovery from decreased serum Ca to its normal range (>9.0 mg/dL) tended to be faster in DFA group (12 h) than in the CONT group (48 h), but the differences were not significant. Active ruminal contraction was observed in DFA group at following parturition of d 1 (p<0.05), d 3 (p<0.05), and d 5 (p<0.01). Dry matter (DM) intake did not differ between the groups. However, positive correlations were observed between serum Ca concentration and ruminal contraction (p<0.001), and between ruminal contraction and DM intake (p<0.001) during following parturition. According to multiple regression analysis (R(2) = 0.824, p<0.001), the DM intake was positively affected by serum Ca concentration and ruminal contraction. These results suggest that feed intake soon after parturition in dairy cows can be increased by improvement of serum Ca concentration and active ruminal contraction, but DFA III supplementation in this study did not improve the lower serum Ca concentration due to parturition.

Donor cell-derived leukemia after cord blood transplantation and a review of the literature: differences between cord blood and BM as the transplant source.

Donor cell-derived leukemia (DCL) is a rare complication of SCT. Here, we present a case of DCL following cord blood transplantation (CBT) and review the clinical features of previously reported DCL. To our knowledge, this is the first report comparing clinical characteristics of DCL from the standpoint of the transplant source, with umbilical cord blood and BM. AML and myelodysplastic syndrome (MDS) were recognized more frequently in DCL after CBT, whereas the incidence of AML and ALL was similar after BMT. The median duration between the occurrence of DCL following CBT and BMT was 14.5 and 36 months, respectively. DCL occurred in a significantly shorter period after CBT than after BMT. Abnormal karyotypes involving chromosome 7 were observed in 52.4% of CBT recipients and 17.3% of BMT recipients; this was a statistically significant difference. Particularly, the frequency of monosomy 7 was significantly higher in DCL after CBT than after BMT. The types of abnormal karyotypes in DCL following BMT were similar to those characteristically observed in adult de novo AML and MDS. DCL patients generally have a poor prognosis in both groups. SCT is the best treatment for curing DCL. DCL appears to have different clinical features according to the transplant source.

Short communication: effect of difructose anhydride III on serum immunoglobulin G concentration in newborn calves.

Difructose anhydride (DFA) III is an indigestible disaccharide that promotes paracellular absorption of calcium, magnesium, and other minerals in the intestine by acting on epithelial tight junctions. This study aimed to elucidate the effect of DFA III on serum IgG concentration. One hundred and twenty Holstein and Holstein/Japanese Black crossbred calves were randomly divided into 4 groups of 30 to receive untreated colostrum (DFA0) or colostrum containing 3, 6, or 18 g of DFA III (DFA3, DFA6, or DFA18, respectively). At 24 h after birth, both serum IgG (ranging from 16.4 to 21.2 mg/mL) and apparent efficiency of absorption (26.0 to 37.2%) showed increases with the amount of DFA III intake. By multiple regression analysis, the standardized partial regression coefficient for DFA III was 0.25, the second highest following that for the colostrum IgG concentration (0.80), indicating a positive effect of DFA III on serum IgG. A positive linear regression was found between colostrum IgG and serum IgG concentrations at 24h of age. These results indicate that IgG absorption occurred as a nonsaturable process, which might be characteristic of gradient-dependent paracellular transport. Thus, it was concluded that DFA III improves not only minerals but IgG absorption in calves.

Non-oxidative glucose disposal is reduced in type 2 diabetes, but can be restored by aerobic exercise.

Whole-body glucose utilization consists of mitochondrial glucose oxidation and non-oxidative glycogen synthesis. We examined whether reduction of both non-oxidative glucose disposal and glucose oxidation contributes to insulin resistance in type 2 diabetes. We also examined the effects of exercise on these two components. Whole-body glucose disposal rate (GDR, mg/kg/min) was evaluated in 37 type 2 diabetic (T2DM) and 17 non-diabetic (non-DM) subjects as the mean of glucose infusion rate during steady state in the euglycaemic-hyperinsulinaemic clamp study. Glucose oxidation rates were assessed by indirect calorimetry, and non-oxidative GDR was calculated by subtracting glucose oxidation rate from GDR. Intramyocellular lipid (IMCL) content of the soleus muscle was measured using (1)H-magnetic resonance spectroscopy. In 10 T2DM subjects, the changes in oxidative and non-oxidative glucose disposal during clamp were examined after 3-month exercise intervention. GDR (2.93 +/- 1.55 vs. 4.55 +/- 1.83, p = 0.001) and non-oxidative GDR (1.45 +/- 1.52 vs. 3.01 +/- 1.87, p = 0.002) were significantly lower in T2DM than in non-DM subjects. Glucose oxidation rate was comparable in the two groups, and inversely correlated with IMCL (n = 15, r =-0.565, p = 0.028). GDR (2.28 +/- 1.67 to 4.63 +/- 2.42, p = 0.021) and non-oxidative GDR (0.72 +/- 1.27 to 2.26 +/- 1.91, p = 0.047) were increased after exercise intervention, although the change in glucose oxidation rate was not significant. In summary, reduction of non-oxidative glucose disposal may contribute to decreased whole-body glucose utilization. In addition, exercise improves insulin resistance mainly by increasing non-oxidative glucose disposal in type 2 diabetes.

Content of reticulocyte hemoglobin is a reliable tool for determining iron deficiency in dialysis patients.

BACKGROUND: The evaluation of iron status in dialysis patients provides information essential to the planning of adequate recombinant human erythropoietin (rHuEPO) treatment. Iron status of the patients can be determined from the recently available measurement of content of reticulocyte hemoglobin (CHr). METHODS: In this study, to clarify the accuracy of CHr in diagnosing iron deficiency in hemodialysis (HD) patients, we initially compared CHr with such conventional iron parameters as serum ferritin levels, transferrin saturation and serum soluble transferrin receptor levels. Secondly, we investigated the changes in CHr during iron supplementation for iron-deficient patients to determine whether this marker is a prospective and reliable indicator of iron sufficiency. The participants in this study were 149 hemodialysis (HD) patients and 53 age-matched healthy subjects. Iron deficiency was defined as having a TSAT of less than 20% and serum ferritin of less than 100 ng/ml. Conventional parameters of red blood cells and CHr were measured by an ADVIA120 autoanalyzer. RESULTS: Mean CHr was 32.3 +/- 2.2 pg in the patients undergoing hemodialysis treatment. CHr significantly correlated with iron parameters in the dialysis patients. Logistic regression analysis was performed to determine the relationship between CHr and each outcome measure, and CHr was the significant multivariate predictor of iron deficiency. Iron supplements given to the patients with low CHr and hematocrit (Hct) significantly increased Hct, resulting in a decrease in the weekly dosage of rHuEPO. CONCLUSIONS: CHr, measured simultaneously with Hct, is a sensitive and specific marker of iron status in dialysis patients.

Relative increase of granulocytes with a paroxysmal nocturnal haemoglobinuria phenotype in aplastic anaemia patients: the high prevalence at diagnosis.

To clarify the pathologic significance of granulocytes exhibiting the paroxysmal nocturnal haemoglobinuria (PNH) phenotype in patients with aplastic anaemia (AA), we examined peripheral blood from 100 patients with AA for the presence of granulocytes deficient in glycosylphosphatidylinositol (GPI)-anchored proteins using a sensitive flow cytometric assay. A significant increase in the frequency of CD55-CD59-CD11b+ granulocytes (>0.003%) compared to normal individuals was observed in 31 of 35 (88.6%) patients with untreated AA at diagnosis. The proportions of patients showing increased PNH granulocytes in treated AA patients with a short (<5 yr) and long (>5 yr) disease duration were 68.6% (11/16) and 20.4% (10/49), respectively. When 19 patients showing increased frequency of PNH granulocytes before therapy were studied 6-12 months after antithymocyte globulin plus cyclosporin A therapy, the frequency decreased to 0.01-90% of pretreatment values in 15 recovering patients. These findings suggest that a relative increase in the number of PNH granulocytes is a common feature of AA at diagnosis, and that it may represent the presence of immunologic pressure to normal haematopoietic stem cells as a cause of AA.

Anti-lymphoma effect of naproxen and indomethacin in a patient with relapsed diffuse large B-cell lymphoma.

A 77-year-old man with relapsed non-Hodgkin's lymphoma, diffuse large B-cell type, was treated with naproxen, a nonsteroidal anti-inflammatory drug (NSAID), for paraneoplastic fever. A dramatic disappearance of not only the fever but also generalized lymphadenopathy was observed. Naproxen was continued, and he maintained remission for 10 months. When relapse of lymphoma occurred in spite of continuous naproxen administration, indomethacin, another type of NSAID, was tried. Surprisingly, rapid regression of lymphoma occurred again and was maintained for almost 1 year. These results indicate that NSAIDs are effective in some patients with non-Hodgkin's lymphoma.

Chemoprotective effects of KF41399, a derivative of carbazole compounds, on nimustine-induced thrombocytopenia.

We examined the chemoprotective effects of KF41399, a novel derivative of carbazole compounds, on severe thrombocytopenia induced by nimustine (ACNU, 45 mg/kg administered for 2 consecutive days intravenously) in mice. Administration schedule studies revealed that pretreatment of mice with KF41399 was necessary to improve thrombocytopenia. Oral administration of KF41399 ameliorated thrombocytopenia induced by ACNU and accelerated the rate of platelet recovery in a dose-dependent fashion. In addition, KF41399 pretreatment improved the decrease in body weight and spleen weight and in the colony-forming activity of bone marrow mononuclear cells (MNC). Oral administration of KF41399 to normal mice induced G(0)/G(1)-phase accumulation of MNC as well as hematopoietic progenitor cells (lineage negative cells [Lin(-)]) and reduced the colony-forming activity of MNC. In Lin(-) cells derived from KF41399-treated mice, up-regulation of Bcl-2 and down-regulation of cyclin E and cyclin A proteins were observed. In the same cells, a decrease in the phosphorylated form of Rb protein and an increase in the p130 protein were observed without changes in the protein level of cell cycle-dependent kinase 2 (Cdk2), Cdk4, and Cdk6. More important, KF41399 did not affect the antitumor activity of ACNU against mouse Sarcoma180 and human lung cancer LC-6. However, 25-mg/kg KF41399 treatment reduced the antitumor activity of ACNU against human lung cancer Lu-65, and 5 mg/kg KF41399 caused a slight reduction of the antitumor activity of ACNU without inducing thrombocytopenia. These results suggest that KF41399 might be useful as a chemoprotective agent to improve chemotherapy-induced thrombocytopenia and types of other toxicity. (Blood. 2000;95:3771-3780)

Delineation of the frequently deleted region on chromosome arm 13q in B-cell non-Hodgkin's lymphoma.

The loss of a specific chromosomal region provides a clue to the elucidation of the putative tumor suppressor gene implicated in the pathogenesis and progression of tumors. To delineate the specific region(s) involved in lymphomagenesis, we performed a survey of loss of heterozygosity for 11 polymorphic microsatellite loci scattered on variable chromosome arms. We examined 20 primary lymphoma samples, including both indolent and aggressive B-cell non-Hodgkin's lymphoma (B-NHL) and Hodgkin's disease (HD), and found a significant number of B-NHLs with loss of genetic material on chromosome arm 13q at the RB1 locus (50%; 4 of 8 informative cases for the RB1 locus). To specify the 13q deletion and to narrow the critical deleted region, we examined the same 20 lymphomas by intensive microsatellite mapping analysis using 12 microsatellite markers, mapping from 13q12.3 to 13q14. We confirmed the frequent 13q14 deletion to be in the vicinity of the RB1 locus (50% of the informative NHLs for at least 1 of 12 microsatellite loci; 5 of 10 aggressive NHLs and 2 of 4 indolent NHLs, but none of 6 HDs) and determined a subchromosomal region deleted in lymphoma on 13q14 defined by D13S164-D13S273, which is an overlapped region frequently lost in chronic lymphocytic leukemia. Taken together, our data indicate that the 13q alterations are present in a wide variety of NHLs including both indolent and aggressive B-NHLs, suggesting that loss of genetic material at chromosome band 13q14 may play an important role in the formation or development of a wide variety of mature lymphoid malignancies.

Vitrectomy for vitreous bleeding and tractional retinal detachment in a case of Evans syndrome.

PURPOSE: To evaluate the effect of vitrectomy to treat Evans syndrome with vitreous bleeding followed by tractional retinal detachment. CASE: Vitrectomy was performed on a 21-year-old man with Evans syndrome who developed tractional retinal detachment in the left eye after experiencing vitreous bleeding and tissue proliferation related to the bleeding. OBSERVATIONS: After the proliferative tissue was removed and tractional retinal detachment corrected, the patient's visual acuity improved from 6/20 to 10/20. A small amount of residual proliferative tissue remained after the vitrectomy. CONCLUSIONS: Vitrectomy may be beneficial in patients with serious vitreous complications induced by hematological diseases such as Evans syndrome.

Frequent chromosome arm 13q deletion in aggressive non-Hodgkin's lymphoma.

To clarify the role of allelic loss on chromosome arm 13q in lymphomagenesis, we performed fluorescence in situ hybridization (FISH) analysis of a total of 43 primary lymphomas, including both indolent and aggressive non-Hodgkin's lymphoma (NHL) and Hodgkin's disease (HD), using the specific probes at RB1 and D13S319 loci on the centromeric portion of chromosome arm 13q. Monosomy at either or both RB1 and D13S319 loci was detected in 15 of 43 (35%) lymphomas (14 of 43 cases at RB1 locus and seven of 43 cases at D13S319 locus); the 13q deletion was frequently detected in the aggressive NHLs (40%; 12 of 30 cases) compared with that in indolent NHL (17%; one of six cases) and a subset of HD (29%; two of seven cases). There are only six cases of 43 which have total monosomy 13q14, all aggressive NHL, 14% of total or 20% of this subgroup. In addition, we analyzed the loss of heterozygosity in 15 of the 43 primary lymphoma samples for several polymorphic microsatellite loci (D13S168, RB1 and D13S272) on the chromosome arm 13q, and confirmed the 13q deletion in four of five cases that were positive on FISH analysis. The subchromosomal region frequently altered in lymphoma on 13q14 is the region around RB1 locus and centromeric to D13S319 locus, which is an overlapped region frequently deleted in chronic lymphocytic leukemia. Together, our data indicate that the 13q alterations are present in a variety of types of lymphoma and occur in a significant proportion of aggressive NHLs, suggesting the possible presence of common candidate gene(s) on the 13q14 region, whose alteration may play an important role in the formation or development of a wide variety of mature lymphoid malignancies.

[Thrombopoietin: current and future status].

Thrombopoietin (TPO), which is a major physiological regulator of platelet production, was cloned in 1994. It has already been shown that the administration of recombinant TPO increases platelet production and accelerates platelet recovery after cytoablative therapy in preclinical and clinical trials. In addition, recent results indicate that the effects of TPO on hematopoiesis are more widespread than initially anticipated. In this article, physiological activities of TPO and results of clinical trials are briefly reviewed, and possible clinical application and some problems in its clinical use are discussed.

In vitro and in vivo effects of KT6352, a derivative of indolocarbazole compounds, on murine megakaryocytopoiesis.

We investigated the in vitro and in vivo effects of KT6352, a derivative of indolocarbazole compound, on murine megakaryocytopoiesis. When serum-free megakaryocyte (Meg) colony assay was performed with 100 U/mL of recombinant mouse interleukin-3 (rmIL-3), the addition of 1x10(-11)M to 1x10(-9)M of KT6352 increased the number of Meg colonies. An additional increase of Meg colonies by KT6352 was observed in the serum-free culture containing rmIL-3 plus recombinant mouse interleukin-6 or rmIL-3 plus recombinant mouse stem cell factor. KT6352 did not stimulate Meg colony formation without rmIL-3. When KT6352 was administered intraperitoneally to normal BALB/c male mice at a dose of 10 mg/kg daily for 5 consecutive days, a 2.1-fold increase in the platelet count was observed on day 14, and the prolonged thrombocytopoiesis was detectable from 9 to 27 days after KT6352 administration. A marked increase in the white blood cell count was also observed from 5 to 14 days after KT6352 treatment. Before the gradual increase of platelet counts, 8 days after KT6352 administration, a marked increase in the number of colony-forming units of megakaryocytes (CFU-Megs) in bone marrow and spleen was observed, and a substantial increase in the number of splenic CFU-Megs was observed 14 and 23 days after KT6352 administration. Bone marrow Meg ploidy analysis by two-color flow cytometry showed a shift in the modal ploidy class from 16 to 32 and an increase in the frequency of 64 cells in KT6352-treated mice. These results suggest a possible therapeutic benefit of KT6352 in the management of thrombocytopenia.

K-252a-induced polyploidization and differentiation of a human megakaryocytic cell line, Meg-J: transient elevation and subsequent suppression of cyclin B1 and cdc2 expression in the process of polyploidization.

Megakaryocytes are unique haemopoietic cells which undergo DNA replication, giving rise to polyploid cells. However, little is known about the mechanism of megakaryocytic polyploidization. To address this issue, we used the human megakaryocytic cell line Meg-J. In the presence of K-252a (an indolocarbasole derivative), Meg-J cells stopped proliferation and exhibited additional megakaryocytic features, including morphological changes, polyploidization, and increases in the levels of surface expression of platelet glycoprotein (GP) IIb/IIa and GPIb. Thrombopoietin (TPO) promoted the K-2 52a-induced polyploidization and megakaryocytic differentiation. In the process of K-252a-induced polyploidization, levels of expression of both cdc2 and cyclin B1 were elevated transiently and subsequently decreased. This suggested that the polyploidization process in Meg-J cells was at least in part associated with a transient elevation and subsequent decrease in the expression of cdc2/cyclin B1 complex, a critical kinase involved in G2/M cell cycle transition.

Transcription factor NF-E2 is essential for the polyploidization of a human megakaryoblastic cell line, Meg-J.

Transcription factors regulating the process of megakaryocyte development remain largely unclarified. To clarify them further, we used a human megakaryoblastic cell line, Meg-J, which showed prominant polyploidization and augmented platelet glycoprotein (GP) Ib expression after incubation with thrombopoietin (TPO, c-mpl ligand) and K252a (an indolocarbasole derivative). Under these conditions, we analyzed the expression of the transcription factors and observed that the expression of NF-E2 p45, but not those of GATA-1, GATA-2, Tal-1/SCL, Evi-1, and MafK, was increased after TPO and K252a stimulation. Gel-shift assay confirmed the enhanced binding activity to the NF-E2 site. The abolishment of NF-E2 p45 with NF-E2 antisense oligomers inhibited TPO plus K252a-induced polyploidization. These findings suggest that NF-E2 p45 is essential for the polyploidization of megakaryocytic cells.