Occurrence of adverse events caused by valganciclovir as pre-emptive therapy for cytomegalovirus infection after allogeneic stem cell transplantation is reduced by low-dose administration.

BACKGROUND: Pre-emptive therapy with valganciclovir (VGCV) has become the standard therapy for preventing cytomegalovirus (CMV) infection after allogeneic hematopoietic stem cell transplantation (HSCT). The effectiveness of low-dose VGCV (900 mg per day) has been shown to be equal to that of standard-dose VGCV (900 mg twice daily); however, individualized optimal dosing and toxicity of VGCV have not been reported. METHODS: We conducted a retrospective study to evaluate the optimal dose of VGCV as pre-emptive therapy for preventing CMV infection by comparing the frequency of adverse events (AEs) and clinical efficacy in a low-dose VGCV group with those in a standard-dose VGCV group. Thirty-eight patients who were administered VGCV because of CMV antigenemia after HSCT were analyzed. RESULTS: Neutropenia (standard-dose group: 33%, low-dose group: 15%, P = 0.26) and thrombocytopenia (standard-dose group: 39%, low-dose group: 15%, P = 0.14) were frequent AEs of VGCV, and a significantly higher frequency of overall AEs was detected in the standard-dose group than in the low-dose group (P < 0.01). In comparison of dosage based on weight, dosage of VGCV >27 mg/kg was closely related to onset of AEs (P = 0.04). CONCLUSIONS: Low-dose VGCV was not inferior in clinical efficacy, including clearance rate of CMV antigenemia and incidence of consequent CMV disease, to standard-dose VGCV as was previously reported. Initial low-dose VGCV for pre-emptive CMV therapy markedly reduces hematologic toxicity and has clinical efficacy equivalent to that of standard-dose VGCV. It is therefore reasonable for patients, except for noticeably overweight patients, to be given initial low-dose VGCV.

Hepatitis B virus (HBV) reverse seroconversion (RS) can be prevented even in non-responders to hepatitis B vaccine after allogeneic stem cell transplantation: long-term analysis of intervention in RS with vaccine for patients with previous HBV infection.

BACKGROUND: Reactivation of hepatitis B virus (HBV) infection, reverse seroconversion (RS), is a serious complication after allogeneic stem cell transplantation (alloHSCT). We previously conducted a post-transplant hepatitis B vaccine intervention trial and demonstrated the vaccine efficacy in preventing HBV-RS. This report is an update of the hepatitis B vaccine study. METHODS: In this trial, 21 patients were enrolled and received a standard 3-dose regimen of hepatitis B vaccine after discontinuation of immunosuppressants, whereas 25 transplant recipients with previous HBV infection did not receive the vaccine and served as controls. RESULTS: None of the 21 patients in the vaccine group developed HBV-RS and 12 controls developed HBV-RS in median follow-up periods of 60 months (range 13-245). HBV vaccine resulted in a positive value of hepatitis B surface antibody (HBsAb) titer in 9 patients, while HBsAb remained negative in 12 patients. Presence of a high titer of HBsAb before vaccination was associated with conversion into HBsAb positivity after vaccination. CONCLUSION: These results demonstrated the long-term effects of HBV vaccine for preventing HBV-RS after alloHSCT. Of note, no HBV-RS occurred, even in patients who did not achieve conversion into HBsAb positivity after vaccination.

Efficacy of folinic acid in preventing oral mucositis in allogeneic hematopoietic stem cell transplant patients receiving MTX as prophylaxis for GVHD.

As the safety of folinic acid administration and its efficacy for reducing the toxicity of MTX remain controversial, we assessed the effect of folinic acid administration after MTX treatment for GVHD prophylaxis on the incidence of oral mucositis and acute GVHD. We retrospectively analyzed data for 118 patients who had undergone allogeneic hematopoietic SCT and had received MTX for GVHD prophylaxis. Multivariate analysis showed that systemic folinic acid administration significantly reduced the incidence of severe oral mucositis (odds ratio (OR)=0.13, 95% confidence interval (CI) 0.04-0.73, P=0.014). There was also a tendency for a lower incidence of severe oral mucositis in patients who received folinic acid mouthwash (OR=0.39, 95%CI 0.15-1.00, P=0.051). No significant difference was observed in the incidence of acute GVHD between patients who received systemic folinic acid administration and those who did not (P=0.88). Systemic folinic acid administration and mouthwash appear to be useful for reducing the incidence of severe oral mucositis in patients who have received allogeneic hematopoietic SCT using MTX as GVHD prophylaxis.

Increased risk of bacterial infection after engraftment in patients treated with allogeneic bone marrow transplantation following reduced-intensity conditioning regimen.

Although bacterial infection is a major cause of death even after reduced-intensity conditioning (RIC) for allogeneic stem cell transplantation (SCT), little is known about the epidemiology and risk factors. The incidence of bacterial infection in 43 patients who received allogeneic bone marrow transplantation (BMT) using a RIC regimen was compared with that in 68 patients who received BMT using a myeloablative conditioning regimen, and risk factors for bacterial infection were identified. Before engraftment, incidences of febrile neutropenia (FN) and documented infections (DI) were significantly decreased in RIC patients (FN: 59.5% vs. 89.6%, P<0.01, DI: 4.8% vs. 17.9%, P<0.01). However, incidence of bacterial infection was significantly increased in RIC patients in the post-engraftment phase (53.8% vs. 11.1%, log-rank, P<0.01). Blood stream was the most frequent focus of infection in both groups. In multivariate analysis, RIC and acute graft-versus-host disease were revealed to be significant risk factors for bacterial infection in this phase. In summary, risk of bacterial infection after engraftment was significantly higher in RIC patients, although infection was decreased before engraftment, and we need to develop a RIC-specific strategy against bacterial infection after RIC SCT.

Chimerism and T-cell receptor repertoire analysis after unrelated cord blood transplantation with a reduced-intensity conditioning regimen following autologous stem cell transplantation for multiple myeloma.

A 65-year-old Japanese male was diagnosed as multiple myeloma with Bence Jones kappa type, clinical stage IIIA. His disease status reached partial remission after chemotherapy. Thereafter, he received tandem transplantation, consisting of high-dose chemotherapy with autologous stem cell transplantation (ASCT), followed by unrelated cord blood transplantation (U-CBT). U-CBT with a reduced-intensity conditioning regimen (RI-CBT) was performed in August 2003. HLA mismatch between the patient and the CBT donor was present at two serological loci (B and DR). A total nucleated CBT cell dose of 2.45 x 10(7)/kg body weight was infused on day 0. Graft-vs.-host disease (GVHD) prophylaxis consisted of cyclosporine A and short-term methotrexate. Neutrophil engraftment (>0.5 x 10(9)/l) was obtained on day 46. He developed positive cytomegalovirus antigenemia, grade II acute GVHD involving skin and liver, varicella-zoster virus infection, septic shock, hemorrhagic cystitis caused by adenovirus and acute hepatitis B virus infection after U-CBT. We retrospectively analyzed T-cell receptor (TCR) repertoire diversity and found that TCR repertoire diversity decreased continuously after U-CBT. Therefore, low-TCR repertoire diversity in this patient appears to be associated with various infections caused by immunodeficiency.

Regulation of the expression of MHC class I-related chain A, B (MICA, MICB) via chromatin remodeling and its impact on the susceptibility of leukemic cells to the cytotoxicity of NKG2D-expressing cells.

Innate immune cells such as natural killer (NK) cells play a crucial role in antitumor immune responses. NKG2D is a major activating immunoreceptor expressed in not only NK cells but also CD8+ T cells and shows cytotoxicity against tumors by recognizing its ligands major histocompatibility complex class I-related chain A and B (MICA and MICB) on tumor cells. Recently, it has been suggested that NKG2D-mediated cytotoxicity correlates with the expression levels of NKG2D ligands on target cells. In this study, we were able to increase the expression levels of MICA and MICB on leukemic cell lines and patients' leukemic cells by treatment with trichostatin A (TsA), a histone deacetylase (HDAC) inhibitor. Chromatin immunoprecipitation (ChIP) assays revealed that treatment with TsA resulted in increased acetylation of histone H3 and decreased association with HDAC1 at the promoters of MICA and MICB. Intriguingly, upregulation of MICA and MICB by treatment with TsA led to enhancement of the susceptibility of leukemic cells to the cytotoxicity of NKG2D-expressing cells. Our results suggest that regulation of the expression of NKG2D ligands by treatment with chromatin-remodeling drugs may be an attractive strategy for immunotherapy.

Analysis of donor-type chimerism in lineage-specific cell populations after allogeneic myeloablative and non-myeloablative stem cell transplantation.

We analyzed donor-type chimerism in CD3+, CD14.15+ and CD56+ cells from 36 patients who had undergone conventional-intensity allogeneic stem cell transplantation (CST) and 34 patients who had undergone non-myeloablative allogeneic stem cell transplantation (NST) for hematological malignancies. On day 28 after transplantation, all fractions in NST patients and CD3+ cells in CST patients who received a non-total body irradiation (TBI) regimen showed more frequent mixed chimerism (<90% donor cells) than those in patients who had received TBI. NST patients with acute graft-versus-host disease (grade II-IV) frequently showed more than 50% donor-type chimerism in CD3+ cells on day 14 (P=0.029). NST patients with <50% donor-type chimerism on day 14 and with <90% donor-type chimerism on day 28 in CD56+ cells had significantly poor 1-year overall survival (0 vs 91%, P<0.001 and 20 vs 74%, P=0.002, respectively). Both NST and CST patients with <90% donor-type chimerism in CD14.15+ cells on day 28 had significantly poor 1-year overall survival (14 vs 70%, P=0.005 and 0 vs 66%, P=0.002, respectively). Our data show that the extent of donor-type chimerism in lineage-specific cells appears to have an impact on outcome after allogeneic stem cell transplantation.

Does carotid artery stenting work on the long run: 5-year results in high-volume centers (ELOCAS Registry).

AIM: Although the first long-term results of Carotid Artery Stenting (CAS) became available only recently, CAS has become an accepted treatment for carotid artery disease. We report CAS data up to 5 years, both late stroke rate and patency rates as observed in 4 high-volume European centers. METHODS: Between February 1, 1993 and December 31, 2004, 2 172 patients were selected over the 4 participating centres, with intention to treat endovascularly. Conscientious follow-up was done according to the in-hospital stipulations of each centre and was entered into a database both retrospectively and prospectively. Long-term restenosis and stroke-death rates were investigated and statistically analysed and stratified using the Kaplan-Meier method. RESULTS: Of the 2 172 patients with intention to treat 2 165 (99.7%) were technically successful. Of these 306 (14.1%) were performed without and 1 859 (85.9%) with embolic protection device (EPD); 96 patients (4.4%) received balloon dilation only and stenting was performed in 2 069 (95.6%) cases. Kaplan-Meier analysis of major stroke/all death and of significant restenosis (>50%) for the total population showed stroke/death rates of 4.1% (nar=1 356), 10.1% (nar=476) and 15.5% (nar=138); and restenosis rates of 1% (nar=1 363), 2% (nar=480) and 3.4% (nar=139), after 1, 3 and 5 years respectively. CONCLUSIONS: The patency and stroke/death rates resulting from our database analysis are pleasing and indicate that CAS also on longer term is a valuable treatment method for carotid artery disease. Due to the fact that our dataset contains prospective as well as retrospective data, it may have its limitations. Until this moment, data indicating that certain patient subgroups are at increased risk for neurological complications and in-stent restenosis during and after CAS are sparse. Further multivariant analysis on this unique dataset is mandatory in order to identify any potential links in between plaque morphology, preprocedural neurological complications, risk factor distribution, procedural steps and clinical outcome.

Expression of Dax-1 during gonadal development of the frog.

Dax-1, a member of the nuclear hormone receptor superfamily of transcription factors, is known to be involved in gonadal development in mammals. To date, Dax-1 has only been isolated in reptiles, birds and mammals. The expression of Dax-1 is down-regulated in the developing testis, but persists in the ovary of mice (Swain et al., Nat. Genet. 12 (1996) 404) and chicken (Smith et al., J. Mol. Endocrinol. 24 (2000) 23). Curiously, there is no sex difference in the expression patterns of Dax-1 in the American alligator (Western et al., Gene 241 (2000) 223). To understand its role(s) in gonadal development in vertebrates, molecular cloning of Dax-1 in amphibians is required. In this study, we cloned an amphibian Dax-1 homologue of the frog Rana rugosa and examined its expression profile during gonadal development. Cloned Dax-1 cDNA encoded a protein of 287 amino acids. Unlike mammalians that possess the three and one half repeat elements representing the putative DNA binding domain in the predicted sequence of Dax-1 protein, the frog had a single poorly conserved copy of the repeat unit. By RT-PCR analysis, the Dax-1 mRNA was detected in the liver and pancreas, but not in the testis and ovary of adult frogs. However, Dax-1 expression was seen first in the embryo at stage 12 and became stronger in tadpoles until stage X. The Dax-1 was transcribed in the testis stronger than in the ovary of frogs at stage XXV (just after completion of metamorphosis). In the gonad of frogs 2 months after metamorphosis (at this stage postmeiotic cells can be seen in the seminiferous tubules), the Dax-1 was expressed only in males. In addition, the Dax-1 transcription declined gradually as ovarian development proceeded, but its expression was down-regulated and then up-regulated rapidly when female-to-male sex reversal was caused by administration of testosterone into female tadpoles. Taken together, the results suggest that the Dax-1 may be closely involved in testicular development of amphibians.

LST-2, a human liver-specific organic anion transporter, determines methotrexate sensitivity in gastrointestinal cancers.

BACKGROUND & AIMS: One approach to the development of targeted cancer chemotherapy exploits increased uptake of the agent into neoplastic cells. In this scenario, higher concentrations of the agent in cancer cells are responsible for differential killing, whereas the low concentration in normal human cells decreases side effects. The aim of this study was to isolate an organic anion transporter that is weak in normal cells, but abundantly expressed in cancer cells, to deliver the anticancer drugs to the cells. METHODS: A human liver complementary DNA (cDNA) library was screened with liver-specific transporter (LST)-1 cDNA as a probe. Northern blot analyses were performed using the isolated cDNA (termed LST-2). An LST-2-specific antibody was raised, and immunohistochemical analyses including immunoelectron microscopy were performed. Xenopus oocyte expression system was used for functional analysis. We also established a permanent cell line that consistently expresses LST-2 to examine the relationship between methotrexate uptake and sensitivity. RESULTS: The isolated cDNA, LST-2, has 79.7% of overall homology with human LST-1. LST-2 exclusively expressed in the liver under normal conditions and its immunoreactivity was highest at the basolateral membrane of the hepatocytes around the central vein. Although its weak expression in the liver, LST-2 is abundantly expressed in the gastric, colon, and pancreatic cancers. On the other hand, the LST-1 was only detected in a hepatic cell line. LST-2 transports methotrexate in a saturable and dose-dependent manner. Furthermore, introduction of the LST-2 gene into mammalian cells potentiates sensitivity to methotrexate. CONCLUSIONS: LST-2 is one of the prime candidate molecules for determining methotrexate sensitivity and may be a good target to deliver anticancer drugs to the gastrointestinal cancers.

The detection of bacteria and bacterial biofilms in punctal plug holes.

PURPOSE: An investigation into bacterial biofilm formation on and in punctal plugs. METHODS: The study involved 21 patients with severe dry eye whose puncta were occluded by the use of punctal plugs. Of these, 15 had Sjögren's syndrome, 3 had non-Sjögren's syndrome, 2 had Stevens-Johnson syndrome, and 1 had graft-versus-host disease. From 17 of the 21 subjects, 18 samples of material were extracted from the holes of the punctal plugs (16 unilateral and 1 bilateral) and were subjected to enrichment culture. Nineteen punctal plugs were removed and processed for electron microscopy: 15 by scanning electron microscopy, and 4 by transmission electron microscopy. RESULTS: Positive cultures were found in 8 of 18 (44%) samples of the material extracted from the holes of punctal plugs. In six of these eight cases (75%) the cultured bacterial species was Staphylococcus epidermidis, whereas in the other two cases (25%) it was S. aureus. In 8 of the 15 punctal plugs examined by scanning electron microscopy and in the material extracted from 1 plug that was examined by transmission electron microscopy, there was clear evidence of bacterial colonization. CONCLUSION: Careful observation of patients with punctal plugs is important. If material accumulates in or on a punctal plug, it may contain bacteria and may form a bacterial biofilm. In these cases, replacement of the plug, clearing of the hole, or an alternative treatment should be considered.

Estimation of anterior nucleus of lens by Scheimpflug image before and after pupil dilatation.

PURPOSE: To compare the accuracy of lens transparency evaluations by Scheimpflug image in the anterior nucleus of the lens before and after pupil dilatation. METHODS: Scheimpflug lens images were recorded in 70 eyes of 38 subjects (age: 28-75 years) before and after pupil dilatation, and light scattering intensity measurements before and after dilatation were compared. RESULTS: There was a significant positive correlation between the light scattering intensity before and after dilatation at the anterior cortex, anterior nucleus, and central clear zone of the lens (r > 0.9, P <.0001). CONCLUSIONS: It is possible to estimate the transparency in the anterior nucleus of the lens from the Scheimpflug image without pupil dilatation. If nuclear type cataracts are regarded as a structural marker of aging in epidemiological studies, measuring the light scattering intensity in the anterior nucleus of the lens without dilatation seems to be a safe, useful, and quantitative method.

Acute conjunctivitis associated with biofilm formation on a punctal plug.

BACKGROUND: Punctal plugs are used for the treatment of tear-deficient type dry eye. We recently examined a case of acute conjunctivitis associated with bacterial biofilm formation on a punctal plug. CASE: A 63-year-old woman diagnosed as having tear-deficient type dry eye came to our hospital with a complaint of soreness in her right eye. Punctal plugs had been inserted into this eye 5 1/2 months previously. On the day of her visit, she presented with acute conjunctivitis. OBSERVATIONS: In biomicroscopical examination, the top of the punctal plug was seen to be covered with a whitish soft material. Microbiological analysis performed on a part of this material was positive for Staphylococcus haemolyticus and Candida tropicalis. Scanning electron microscopy of the removed punctal plug revealed widespread bacterial colonization embedded within an extensive extracellular matrix. Treatment consisted of the replacement of the plug, and administration of a combination of antibacterial eyedrops and preservative-free artificial solution. As a result, the acute conjunctivitis cleared up within 1 month. CONCLUSIONS: This case suggests that a punctal plug poses a potential risk of causing the formation of bacterial biofilm. In such a case, replacement of the plug and/or removal of the accumulated materials should be considered.

Tear meniscus changes during cotton thread and Schirmer testing.

PURPOSE: To elucidate the effect of the cotton thread test (CT-T) and Schirmer test (S-T) on the tear reservoir by evaluating the radius of tear meniscus curvature. METHODS: The radii (R) of the central lower tear menisci were measured by a newly developed video meniscometer in 11 eyes of 11 normal volunteers (6 men, 5 women; mean age, 27.7 +/- 3.6 years [SD]) and 9 eyes of 9 patients with tear deficiency and severe dry eye in whom the puncta had been therapeutically occluded (9 women; mean age, 50.6 +/- 10.4 years). In this dry eye group, the absence of reflex tearing, coupled with the absence of lacrimal drainage due to punctal occlusion allowed more precise observation of the removal of tears from the meniscus. A 1-minute CT-T was performed, followed after an interval of 10 minutes by a 1-minute S-T. Tear meniscus curvature was documented before (R:(0)) and during the tests at 30 seconds (R(30)) and 60 seconds (R:(60)). RESULTS: In the normal group, respective R values (CT-T; S-T; mean +/- SD mm) were R(0) (0.26 +/- 0.11; 0.26 +/- 0. 07), R(30) (0.27 +/- 0.16; 0.20 +/- 0.13), and R(60) (0.29 +/- 0.15; 0.23 +/- 0.21); and in the dry eye group, respective R: values (CT-T; S-T) were R(0) (0.59 +/- 0.23; 0.51 +/- 0.19), R(30) (0.52 +/- 0.25; 0.22 +/- 0.09), and R(60) (0.51 +/- 0.19; 0.21 +/- 0.08). It was demonstrated in the dry eye group that R was diminished more by the S-T than by the CT-T in the time course of the measurement (P = 0.01). In the dry eye group alteration of R occurred within the first 30 seconds, and in this group significant correlation was found between R(0) and the S-T result (r = 0.67; P = 0.05), and between R(60)- R(0) and the S-T result (r = -0.81; P = 0.01). Also, there was a significant correlation between R(60)- R(0) and the S-T result in the normal group (r = 0.71; P = 0.02). There were no significant correlations between R(0) or R(60)- R(0) and the CT-T results in either group. CONCLUSIONS: These studies afford some insight into the dynamics of the Schirmer test, suggesting that wetting is influenced by the negative hydrostatic pressure within the tear meniscus. With the protocol used, no conclusion could be drawn about the relation between meniscus radius and wetting of the cotton thread.

Observation of corneal epithelial disturbance through soft contact lens using fluorescein dextran.

PURPOSE: To confirm the usefulness of fluorescein dextran as a vital dye for observing corneal epithelial disturbance through a medical-use soft contact lens. METHODS: Ten percent fluorescein isothiocyanate-dextran (FITC-dextran) was used as the staining dye. Patients with corneal epithelial disturbance who were wearing therapeutic soft contact lenses were observed using FITC-dextran. RESULTS: Through the soft contact lens, FITC-dextran enabled corneal disturbance observation similar to that seen with sodium fluorescein. In a case of corneal perforation, as well, positive Seidel's test was confirmed through the soft contact lens using this technique. CONCLUSION: FITC-dextran is considered a useful dye for examining the corneal epithelial condition without contact lens removal.

[Comparison of primary and secondary Sjögren's syndrome].

PURPOSE: We studied the difference in severity between primary and secondary Sjögren's syndrome (SS). SUBJECTS AND METHODS: Two groups of patients (all females, mean age: 58 years), 31 with primary SS and 18 with secondary SS were studied. We performed the following dry eye tests: fluorescein score and Rose Bengal staining, grading of tear lipid layer interference patterns, measurement of fluorescein break up time, cotton thread test, and Schirmer-I test. Auto antibodies were also investigated. RESULTS: There was no significant difference between primary and secondary SS with respect to any dry eye tests or auto antibodies. In primary SS, however, the presence of anti SS-A antibody was significantly correlated with Rose Bengal scores (p = 0.044). CONCLUSION: The severity of SS is independent of the primary or secondary type. In primary SS, the presence of anti SS-A antibody may be correlated with the severity.