Improvements of Retinal Sensitivity after Intravitreal Injection of Aflibercept in Eyes with Neovascular Age-Related Macular Degeneration with or without Polypoidal Choroidal Vasculopathy.

PURPOSE: We aim to determine the effects of intravitreal aflibercept (IVA) on the mean sensitivity (MS) of the central retina, best-corrected visual acuity (BCVA), and central foveal thickness (CFT) in eyes with neovascular age-related macular degeneration (nAMD) with or without polypoidal choroidal vasculopathy (PCV). METHODS: This was a prospective, interventional study. All eyes were treatment-naive with nAMD with or without PCV. Each eye received 3 monthly IVA injections followed by an IVA injection every 2 months for 12 months. The primary outcome was the change in the MS within the central 2°. The secondary outcomes were the changes in BCVA, CFT, greatest linear dimension (GLD), and percentage of eyes with a dry macula. RESULTS: Thirty-seven eyes of 37 patients were studied. A significant improvement of the MS (dB) was observed +4.9 ± 4.6 dB (mean ± standard deviation) at 3 M (p < 0.001), +5.5 ± 4.9 dB at 6 (p < 0.001), and +7.0 ± 3.4 dB at 12 M (p < 0.001) compared to the baseline in all eyes. The MS of the eyes with non-PCV was not significantly different from that of eyes with PCV (p = 1.00, 1.00, 1.00, and 0.76 at baseline, 3, 6, and 12 M, respectively). The MS of 11 patients whose BCVA remained unchanged was significantly improved by +6.5 ± 2.8 dB at 3 M (p < 0.001), +6.1 ± 4.3 dB at 6 M (p < 0.001), and +6.4 ± 4.8 dB at 12 M (p = 0.003) compared to the baseline. The mean BCVA was significantly improved from the baseline to 3 M (p < 0.001), 6 M (p = 0.027), and 12 M (p = 0.003) in all eyes. The BCVA was improved or maintained in 97% of the patients at 12 M. The mean CFT and GLD were significantly reduced at 12 M (p < 0.001). Twenty-two eyes (71%) had a dry macula at 12 M. CONCLUSIONS: IVA administered by a fixed dosing regimen led to significant improvements of the central MS, BCVA, and macular morphology at 1 year in eyes with nAMD with or without PCV. These results were not significantly different between eyes with non-PCV and with PCV. The improvements of the MS of the retina of the central 2° in a subgroup whose BCVA remained unchanged through the 12-month experimental period was also significant. We conclude that the MS of the central 2° might be a better marker than the BCVA in determining the effectiveness of IVA treatments and might be helpful in determining early effects on the retina before BCVA changes can be detected.

Nickel-chelatase activity of SirB variants mimicking the His arrangement in the naturally occurring nickel-chelatase CfbA.

Metal-tetrapyrrole cofactors are involved in multiple cellular functions, and chelatases are key enzymes for the biosynthesis of these cofactors. CfbA is an ancestral, homodimeric-type class II chelatase which is able to use not only Ni2+ as a physiological metal substrate, but also Co2+ as a nonphysiological substrate with higher activity than for Ni2+. The Ni/Co-chelatase function found in CfbA is also observed in SirB, a descendant, monomeric-type class II chelatase. This is despite the distinct active site structure of CfbA and SirB; specifically, CfbA shows a unique four His residue arrangement, unlike other monomeric class II chelatases such as SirB. Herein, we studied the Ni-chelatase activity of SirB variants R134H, L200H, and R134H/L200H, the latter of which mimics the His alignment of CfbA. Our results showed that the SirB R134H variant exhibited the highest Ni-chelatase activity among the SirB enzymes, which in turn suggests that the position of His134 could be more important for the Ni-chelatase activity than that of His200. The SirB R134H/L200H variant showed lower activity than R134H, despite the four His residues found in SirB R134H/L200H. CD spectroscopy showed secondary structure denaturation and a slight difficulty in Ni-binding of SirB R134H/L200H, which may be related to its lower activity. Finally, a docking simulation suggested that the His134 of the SirB R134H variant could function as a base catalyst for the Ni-chelatase reaction in a class II chelatase architecture.

A case of Hirschsprung's disease with segmental dilatation of the colon.

BACKGROUND: Segmental dilatation of the colon (SDC) is a rare disease that is characterized by an abrupt segment of dilated colon between regions of normal-sized colon. We herein report a case of SDC associated with Hirschsprung's disease (HD). CASE PRESENTATION: The patient developed abdominal distension soon after birth, and enema examination showed localized intestinal dilatation from the descending colon to the sigmoid colon with significant caliber changes on both the oral and anal sides of the dilated colon. The findings of the rectal mucosal biopsy were consistent with HD. We considered this case to be a combination of HD and SDC and performed laparoscopic-assisted Soave pull-through with resection of the dilated colon when the patient was 7 months old. Resected specimens showed steep caliber changes on the oral and anal sides of the dilated colon. In the pathological examination, no ganglion cells were found in the submucosa on the anal side of the dilated colon. Based on the above findings, we finally made the diagnosis of HD with SDC. CONCLUSION: In HD with a characteristic dilated colon, the possibility of SDC should be considered.

Decreased pregnancy rate per embryo transfer in women undergoing assisted reproductive technology after abdominal trachelectomy: A retrospective study.

OBJECTIVE: Abdominal trachelectomy (AT) is a fertility-preservation surgery for patients with early-stage cervical cancer. Few studies have reported the outcomes of assisted reproductive technology (ART) in patients after AT. The aim of this study was to evaluate the outcomes of ART after AT. STUDY DESIGN: In this retrospective study, we compared the ART outcomes of 13 patients who underwent AT at another hospital prior to undergoing ART at our clinic (T group) and 52 control patients (non-T group) who did not undergo AT prior to ART, selected on the basis of age, time of treatment onset, and serum anti-Müllerian hormone concentrations, matched 1:4, respectively. RESULTS: Cumulative live birth rates were 62% (8/13) and 65% (34/52) in the T and non-T groups, respectively (p = 0.795). The total number of oocyte retrieval cycles was 34 in the T group and 95 in the non-T group. In all oocyte retrieval cycles, no significant differences were noted in the number of oocyte retrievals, rate of fertilization, and presence of good-quality blastocysts (Gardner classification ≥ BB). The total number of embryo transfer (ET) cycles was 55 in the T group and 109 in the non-T group. The pregnancy and live birth rates per ET were lower in the T group than those in the non-T group (pregnancy rate, 20% vs. 39%, p = 0.017; live birth rate, 15% vs. 30%, p = 0.028; respectively). Endometrial thickness before ET was lower in the T group vs. the non-T group: median (range): 7.4 (3.5-14.3) mm vs. 9.0 (5.5-14.9) mm, respectively; p < 0.0001. Multivariate logistic regression models showed that age at oocyte retrieval (adjusted odds ratio [OR], 0.76; 95% confidence interval [CI], 0.66-0.87), use of good-quality blastocysts (adjusted OR, 3.23; 95% CI, 1.20-8.67), and history of AT (adjusted OR, 0.28; 95% CI, 0.11-0.72) were associated with the pregnancy rate per ET. CONCLUSION: The pregnancy rate per ET was lower in patients with vs. without a history of AT. Clinicians should be aware of the longer time to pregnancy in patients who undergo ART after AT.

Cycloserine enantiomers inhibit PLP-dependent cysteine desulfurase SufS via distinct mechanisms.

The cysteine desulfurase SufS is a pyridoxal-5'-phosphate-dependent enzyme and is essential for the SUF system, which participates in iron-sulfur cluster biosynthesis. Inhibition of SufS in the SUF system by D-cycloserine (DCS) in Plasmodium falciparum apicoplast has recently been reported, indicating that SufS could be a target for malaria therapeutics. However, the mechanistic details underlying the inhibition of SufS by DCS have not yet been clarified. Moreover, inhibition of SufS by the other enantiomer, L-cycloserine (LCS), has not been investigated. Herein, we investigated the structure-based inhibition mechanisms of SufS by DCS and LCS using Bacillus subtilis SufS, whose catalytic mechanism has been well characterized in comparison to that of the P. falciparum SufS. Surprisingly, DCS- and LCS-mediated inhibitions of SufS occur via distinct mechanisms resulting in pyridoxamine-5'-phosphate (PMP) in DCS-mediated inhibition and PMP-3-hydroxyisoxazole adduct (PMP-isoxazole) in LCS-mediated inhibition. Biochemical and structural evaluation of SufS variants identified conserved His and Arg residues at the active site as the key determinants of the distinct inhibition mechanisms. The importance of structural elements involved in DCS and LCS-mediated inhibitions of SufS provides valuable insights for the structure-based design of new drugs targeting SufS. DATABASE: Structural data are available in PDB database under the accession numbers 6KFY, 7CEO, 7CEP, 7CEQ, 7CER, 7CES, 7CET, 7CEU, 7E6A, 7E6B, 7E6C, 7E6D, 7E6E, and 7E6F.

The nickel-sirohydrochlorin formation mechanism of the ancestral class II chelatase CfbA in coenzyme F430 biosynthesis.

The class II chelatase CfbA catalyzes Ni2+ insertion into sirohydrochlorin (SHC) to yield the product nickel-sirohydrochlorin (Ni-SHC) during coenzyme F430 biosynthesis. CfbA is an important ancestor of all the class II chelatase family of enzymes, including SirB and CbiK/CbiX, functioning not only as a nickel-chelatase, but also as a cobalt-chelatase in vitro. Thus, CfbA is a key enzyme in terms of diversity and evolution of the chelatases catalyzing formation of metal-SHC-type of cofactors. However, the reaction mechanism of CfbA with Ni2+ and Co2+ remains elusive. To understand the structural basis of the underlying mechanisms and evolutionary aspects of the class II chelatases, X-ray crystal structures of Methanocaldococcus jannaschii wild-type CfbA with various ligands, including SHC, Ni2+, Ni-SHC, and Co2+ were determined. Further, X-ray crystallographic snapshot analysis captured a unique Ni2+-SHC-His intermediate complex and Co-SHC-bound CfbA, which resulted from a more rapid chelatase reaction for Co2+ than Ni2+. Meanwhile, an in vitro activity assay confirmed the different reaction rates for Ni2+ and Co2+ by CfbA. Based on these structural and functional analyses, the following substrate-SHC-assisted Ni2+ insertion catalytic mechanism was proposed: Ni2+ insertion to SHC is promoted by the support of an acetate side chain of SHC.

A rare case of extraluminally pedunculated gastrointestinal stromal tumor with postoperative metastasis to pancreas.

The gastrointestinal stromal tumor (GIST) is the most common type of sarcomatous tumor of the gastrointestinal tract. Many GISTs appear as submucosal tumors with intraluminal protrusion. GISTs with malignant features have a high risk of postoperative recurrence or metastasis, usually to the liver or peritoneum. We present a case of gastric GIST with double rarities: arising completely extraluminally with a pedicle and postoperative metastasis to the pancreas. A woman in her seventies diagnosed with a large extraluminal gastric GIST underwent complete removal of the tumor. Nine months later, a solitary metastatic tumor in the pancreas was detected. Imatinib controlled metastasis well for four years before the tumor became resistant. The patient then had a partial pancreatectomy with splenectomy. She is currently free from recurrence. We genetically analyzed the primary and metastatic GISTs and found known mutations related to poor prognosis and imatinib resistance.

Snapshots of PLP-substrate and PLP-product external aldimines as intermediates in two types of cysteine desulfurase enzymes.

Cysteine desulfurase enzymes catalyze sulfur mobilization from l-cysteine to sulfur-containing biomolecules such as iron-sulfur (Fe-S) clusters and thio-tRNAs. The enzymes utilize the cofactor pyridoxal-5'-phosphate (PLP), which forms the external substrate- and product-aldimines and ketimines during catalysis and are grouped into two types (I and II) based on their different catalytic loops. To clarify the structure-based catalytic mechanisms for each group, we determined the structures of the external substrate- and product-aldimines as catalytic intermediates of NifS (type I) and SufS (type II) that are involved in Fe-S cluster biosynthesis using X-ray crystallographic snapshot analysis. As a common intermediate structure, the thiol group of the PLP-l-cysteine external aldimine is stabilized by the conserved histidine adjacent to PLP through a polar interaction. This interaction makes the thiol group orientated for subsequent nucleophilic attack by a conserved cysteine residue on the catalytic loop in the state of PLP-l-cysteine ketimine, which is formed from the PLP-l-cysteine aldimine. Unlike the intermediates, structural changes of the loops were different between the type I and II enzymes. In the type I enzyme, conformational and topological change of the loop is necessary for nucleophilic attack by the cysteine. In contrast, the loop in type II cysteine desulfurase enzymes showed no large conformational change; rather, it might possibly orient the thiol group of the catalytic cysteine for nucleophilic attack toward PLP-l-cysteine. The present structures allow a revision of the catalytic mechanism and may provide a clue for consideration of enzyme function, structural diversity, and evolution of cysteine desulfurase enzymes. DATABASE: Structural data are available in PDB database under the accession numbers 5WT2, 5WT4, 5ZSP, 5ZST, 5ZS9, 5ZSK, 5ZSO, 6KFZ, 6KG0, and 6KG1.

Effects of Akt signaling on nuclear reprogramming.

Reprogramming of the epigenetic state from differentiated to pluripotent cells can be attained by cell fusion of differentiated somatic cells with embryonic stem (ES) cells or transfer of the nucleus of a differentiated cell into an enucleated oocyte. Activation of Akt signaling is sufficient to maintain pluripotency of ES cells and promotes derivation of embryonic germ (EG) cells from primordial germ cells (PGCs). Here we analyzed the effects of Akt signaling on somatic cell nuclear reprogramming after cell fusion and nuclear transfer. We found that forced activation of Akt signaling stimulated reprogramming after cell fusion of ES cells with thymocytes or mouse embryonic fibroblasts. These hybrid cells showed ES cell characteristics, including in vitro and in vivo differentiation capacity. In contrast, Akt signaling significantly reduced the efficiency of reprogramming with nuclear transfer. Our results demonstrate that Akt signaling plays important roles on the nuclear reprogramming of somatic cells.

Gene expression profiling of diffuse large B-cell lymphoma supervised by CD21 expression.

This study investigated the gene expression profiles of 40 cases of diffuse large B-cell lymphoma (DLBCL) according to CD21 expression, a favourable prognostic factor in DLBCL. Signature genes were analysed by Gene Ontology Tree Machine, and genes concerned with the immune system and related categories were significantly upregulated in CD21- DLBCLs. Of 40 DLBCLs, four were germinal centre B cell-like (GCB) and 36 non-GCB. Of the 36 non-GCB DLBCLs, 14 CD21+ DLBCLs showed significantly better overall survival than the 22 CD21- DLBCLs (P = 0.036). Hierarchical cluster analysis of signature genes related to CD21 was applied to previously published data sets, resulting in two groups for each data set, CD21+ type DLBCLs and CD21- type DLBCLs. Survival of CD21+ type DLBCLs was significantly better than that of CD21- type (P = 0.006 and P = 0.004, respectively). In both data sets, CD21+ type DLBCLs predominantly included GCB DLBCLs compared with CD21- type. The top classifier gene of CD21 expression was IGHM, and the five of nine Gene Ontology categories significant in CD21- DLBCLs included IGHM. Immunohistochemical analysis of 216 DLBCLs confirmed that overall survival of surface (s) IgM+ DLBCLs was significantly poorer than that of sIgM- DLBCLs (P = 0.013).

Clinicopathologic significance of loss of CD19 expression in diffuse large B-cell lymphoma.

To clarify the clinicopathologic significance of a loss of CD19 expression in diffuse large B-cell lymphoma (DLBCL), we evaluated CD19 expression immunohistochemically in frozen sections from 227 patients who had received diagnoses of DLBCL according to the World Health Organization classification between 1987 and 2002. Histopathologic features of patients with CD19- DLBCL were reviewed, and their clinical features, immunophenotypes, and prognoses were compared retrospectively with respect to CD19 expression. CD19 expression was positive in 205 patients (90%). The 22 patients with CD19- DLBCL had a median age of 63 years, and the male-female ratio was 11:11. Compared with patients with CD19+ DLBCL, those with CD19- DLBCL more frequently showed elevated lactate dehydrogenase (LDH) levels (73%, P= .011). Morphologically, 15 (79%) of the 19 CD19- DLBCL patients examined showed plasmablastic/plasmacytoid differentiation. Patients with CD19- DLBCL expressed BCL2 protein less frequently than CD19+ DLBCL (P= .042). Especially noteworthy is that half of the patients with CD19- DLBCL died within 2 years after diagnosis. The CD19- DLBCL group showed a survival curve significantly inferior to that for the CD19+ group (P= .034, generalized Wilcoxon test). Our findings demonstrate that loss of CD19 expression in DLBCL is associated with elevated serum LDH levels and a poor prognosis, especially during the early follow-up period.

Microarray reveals differences in both tumors and vascular specific gene expression in de novo CD5+ and CD5- diffuse large B-cell lymphomas.

Malignant lymphoma is a heterogeneous disease with different clinical features. Among diffuse large B-cell lymphomas (DLBCLs), a unique subtype has been identified recently based on cell surface marker CD5 and clinicopathological features. These de novo CD5(+) DLBCLs account for approximately 10% of all of the DLBCLs and have poorer prognosis. To additionally understand this subtype of DLBCLs at the molecular level and to find genes that are differentially expressed in de novo CD5(+) DLBCLs, CD5(-) DLBCLs, and mantle cell lymphomas, which also have poor prognosis, we performed gene expression profiling using cDNA microarray technology. Data from a total of 9 samples of CD5(-) DLBCLs, 11 samples of de novo CD5(+) DLBCLs, and 10 samples of mantle cell lymphomas were acquired. A series of genes were identified that distinguish these three types of lymphomas. Among DLBCL cases, integrin beta1 and/or CD36 adhesion molecules were overexpressed in most cases of CD5(+) DLBCL. An immunohistochemical confirmation study revealed that integrin beta1 was expressed on lymphoma cells, which may account for the high extranodal involvement and poor prognosis of CD5(+) DLBCLs. In contrast, CD36 was overexpressed on vascular endothelia in CD5(+) DLBCLs, although there was no difference in vascularity detected by von Wilbrand factor antibody between CD5(+) and CD5(-) DLBCLs. Those results suggest that CD5(+) and CD5(-) DLBCLs have different gene expression signatures in both tumor cells and their vascular systems.

Haemostatic abnormalities and thrombotic disorders in malignant lymphoma.

We examined haemostatic abnormalities and thrombotic disorders in 217 patients with malignant lymphoma. Plasma levels of fibrinogen and D-dimer were significantly higher in patients with malignant lymphoma than in healthy subjects. The incidence of severe complications, such as disseminated intravascular coagulation (DIC) and interstitial pneumonia (IP), differed with each clinical stage or histological type, but they occurred frequently in stage IV or natural killer (NK) cell lymphoma. Plasma levels of fibrinogen degradation products (FDP) and D-dimer, leukocyte tissue factor (TF) mRNA and plasma TF antigen were significantly higher in stage IV than in stage I, II or III. Plasma levels of FDP, D-dimer, and leukocyte TF mRNA in NK cell lymphoma were markedly higher than in other types of lymphoma. Immunohistochemical staining of NK cell lymphoma revealed that granulocyte macrophage colony-stimulating factor was positive in tumour cells, whereas von Willebrand factor and TF were positive in vascular endothelial cells of surrounding tissue. Our results suggested that patients with stage IV disease and NK cell lymphoma were in abnormal thrombotic and haemostatic state, and may frequently develop DIC and IP. One of the mechanisms of DIC and IP may involve elevated cytokine production by lymphoma cells, which can stimulate the expression of TF in blood cells or surrounding tissue.

[Diffuse large B-cell lymphoma of the knee occurring 70 years after tuberculous arthritis].

A 72-year-old woman presented with a progressive swollen and painful left knee over a period of a month. The patient had a history of tuberculous arthritis of the left knee joint at the age of 1-2 years. Since then the knee showed a deformity and contraction. MRI scan demonstrated a soft tissue tumor measuring 7 x 8 x 6 cm in the front of the knee, a biopsy specimen taken from which revealed large B-cell lymphoma. The tumor cells showed a phenotype of CD3- CD5- CD10- CD20+ CD21- BCL2+, and surface immunoglobulin M-kappa. Since the lymphoma cells were found to be negative for Epstein-Barr virus encoded RNA by means of in situ hybridization, the DLBCL in this patient was thought to be caused by a different mechanism from pyothorax-associated lymphoma. The patient was treated with chemotherapy (CHOP) followed by radiotherapy, and obtained partial remission. There is no evidence of progression seven months after diagnosis.

De novo CD5-positive diffuse large B-cell lymphoma of the temporal bone presenting with an external auditory canal tumor.

We report a 74-year-old woman with primary CD5-positive diffuse large B-cell lymphoma (DLBCL) of the temporal bone. The patient was admitted because of a mass in the left external auditory canal. She was treated with eight courses of CEOP therapy (rituximab was added from the sixth course) followed by radiotherapy of 40 Gy, and complete remission was achieved. The occurrence of malignant lymphoma in the temporal bone, which is an extremely unusual site, may have depended on the peculiarity of CD5-positive DLBCL.

CD21S antigen expression in tumour cells of diffuse large B-cell lymphomas is an independent prognostic factor indicating better overall survival.

To evaluate the clinical significance of CD21S expression of diffuse large B-cell lymphoma (DLBCL) tumour cells, we compared their clinical features, immunophenotype, response to therapy and outcome in relation to CD21S expression. Between 1987 and 1999, frozen sections from 240 DLBCL cases were examined for CD21S expression by immunohistochemical methods. CD21S expression was detected on the tumour cells of 87 (36%) cases. The median age of the CD21S(+) DLBCL cases was 65 years (range: 17-84 years), the male-female ratio was 42:45, and they showed the following clinical features: Eastern Cooperative Oncology Group score >1 in 14%, lactate dehydrogenase greater than normal levels in 38%, extranodal sites >1 in 14%, stages III/IV disease at diagnosis in 29%, B symptoms in 17%, and a high/high-intermediate International Prognostic Index (IPI) in 23%. They also showed a better overall survival (P = 0.00001, log-rank test) and a better complete remission rate (P = 0.00004, chi-square test) than CD21S(-) DLBCL. Moreover, CD21S(+) DLBCL showed a better survival than CD21S(-) DLBCL for both low/low-intermediate and high/high-intermediate risk categories of IPI (P = 0.045 and P = 0.0016 respectively). Multivariate analysis identified CD21S expression as an independent factor for survival when compared with the five IPI factors. These findings indicate that CD21S expression of DLBCL tumour cells is a useful prognostic factor for survival.