Routine Eye Screening by an Ophthalmologist Is Clinically Useful for HIV-1-Infected Patients with CD4 Count Less than 200 /μL

Objective

To investigate whether routine eye screening by an ophthalmologist in patients with HIV-1 infection is clinically useful.

Methods

A single-center, retrospective study in Tokyo, Japan. HIV-1-infected patients aged over 17 years who visited our clinic for the first time between January 2004 and December 2013 and underwent full ophthalmologic examination were enrolled. At our clinic, ophthalmologic examination, including dilated retinal examination by indirect ophthalmoscopy was routinely conducted by ophthalmologists on the first visit. The prevalence of ophthalmologic diseases and associated factors including the existence of ocular symptoms were analyzed.

Results

Of the 1,515 study patients, cytomegalovirus retinitis (CMV-R) was diagnosed in 24 (2%) patients, HIV retinopathy (HIV-R) in 127 (8%), cataract in 31 (2%), ocular syphilis in 4 (0.3%), and uveitis with unknown cause in 8 (0.5%). Other ocular diseases were diagnosed in 14 patients. The CD4 count was <200 /μL in all CMV-R cases and 87% of HIV-R. The prevalence of any ocular diseases, CMV-R, and HIV-R in patients with CD4 <200 /μL were 22%, 3%, and 15%, respectively, whereas for those with CD4 ≥200 /μL were 5%, 0%, and 2%, respectively. No ocular symptoms were reported by 71% of CMV-R cases and 82% of patients with any ocular diseases.

Conclusions

Routine ophthalmologic screening is recommended for HIV-1-infected patients with CD4 <200 /μL in resource-rich settings based on the high prevalence of ocular diseases within this CD4 count category and because most patients with ocular diseases, including those with CMV-R, were free of ocular symptoms.     

Comparison of the Japanese Orthopaedic Association (JOA) Score and Modified JOA (mJOA) Score for the Assessment of Cervical Myelopathy: A Multicenter Observational Study

Objectives

The Japanese Orthopaedic Association (JOA) score is widely used to assess the severity of clinical symptoms in patients with cervical compressive myelopathy, particularly in East Asian countries. In contrast, modified versions of the JOA score are currently accepted as the standard tool for assessment in Western countries. The objective of the present study is to compare these scales and clarify their differences and interchangeability and verify their validity by comparing them to other outcome measures.

Materials and Methods

Five institutions participated in this prospective multicenter observational study. The JOA and modified JOA (mJOA) proposed by Benzel were recorded preoperatively and at three months postoperatively in patients with cervical compressive myelopathy who underwent decompression surgery. Patient reported outcome (PRO) measures, including Japanese Orthopaedic Association Cervical Myelopathy Evaluation Questionnaire (JOACMEQ), the Short Form-12 (SF-12) and the Neck Disability Index (NDI), were also recorded. The preoperative JOA score and mJOA score were compared to each other and the PRO values. A Bland-Altman analysis was performed to investigate their limits of agreement.

Results

A total of ninety-two patients were included. The correlation coefficient (Spearman’s rho) between the JOA and mJOA was 0.87. In contrast, the correlations between JOA/mJOA and the other PRO values were moderate (|rho| = 0.03 – 0.51). The correlation coefficient of the recovery rate between the JOA and mJOA was 0.75. The Bland-Altman analyses showed that limits of agreement were 3.6 to -1.2 for the total score, and 55.1% to -68.8% for the recovery rates.

Conclusions

In the present study, the JOA score and the mJOA score showed good correlation with each other in terms of their total scores and recovery rates. Previous studies using the JOA can be interpreted based on the mJOA; however it is not ideal to use them interchangeably. The validity of both scores was demonstrated by comparing these values to the PRO values.     

Identification and Characterization of a Novel Galactofuranose-Specific β-D-Galactofuranosidase from Streptomyces Species

β-D-galactofuranose (Galf) is a component of polysaccharides and glycoconjugates and its transferase has been well analyzed. However, no β-D-galactofuranosidase (Galf-ase) gene has been identified in any organism. To search for a Galf-ase gene we screened soil samples and discovered a strain, identified as a Streptomyces species by the 16S ribosomal RNA gene analysis, that exhibits Galf-ase activity for 4-nitrophenyl β-D-galactofuranoside (pNP-β-D-Galf) in culture supernatants. By draft genome sequencing of the strain, named JHA19, we found four candidate genes encoding Galf-ases. Using recombinant proteins expressed in Escherichia coli, we found that three out of four candidates displayed the activity of not only Galf-ase but also α-L-arabinofuranosidase (Araf-ase), whereas the other one showed only the Galf-ase activity. This novel Galf-specific hydrolase is encoded by ORF1110 and has an optimum pH of 5.5 and a Km of 4.4 mM for the substrate pNP-β-D-Galf. In addition, this enzyme was able to release galactose residue from galactomannan prepared from the filamentous fungus Aspergillus fumigatus, suggesting that natural polysaccharides could be also substrates. By the BLAST search using the amino acid sequence of ORF1110 Galf-ase, we found that there are homolog genes in both prokaryotes and eukaryotes, indicating that Galf-specific Galf-ases widely exist in microorganisms.     

Pharmacological assessment of methamphetamine-induced behavioral hyperactivity mediated by dopaminergic transmission in planarian Dugesia japonica

The freshwater planarian Dugesia japonica has a simple central nervous system (CNS) and can regenerate complete organs, even a functional brain. Recent studies demonstrated that there is a great variety of neuronal-related genes, specifically expressed in several domains of the planarian brain. We identified a planarian dat gene, named it D. japonica dopamine transporter (Djdat), and analyzed its expression and function. Both in situ hybridization and immunofluorescence revealed that localization of Djdat mRNA and protein was the same as that of D. japonica tyrosine hydroxylase (DjTH). Although, dopamine (DA) content in Djdat(RNAi) planarians was not altered, Djdat(RNAi) planarians showed increased spontaneous locomotion. The hyperactivity in the Djdat(RNAi) planarians was significantly suppressed by SCH23390 or sulpiride pretreatment, which are D₁ or D₂ receptor antagonists, respectively. These results suggest that planarians have a Djdat ortholog and the ability to regulate dopaminergic neurotransmission and association with spontaneous locomotion.     

Characterization of abdominal appendages in the sawfly, Athalia rosae (Hymenoptera), by morphological and gene expression analyses

Larvae of the sawfly, Athalia rosae, have remarkable abdominal prolegs. We analyzed the morphogenesis of appendages and the expression of decapentaplegic and Distal-less genes during embryonic development to characterize the origin of prolegs. Proleg primordia in abdominal segments A1–A9 appeared shortly after the inner lobes (endites) of gnathal appendages were formed. These were located on the ventral plates, medioventral to the appendages of the other segments in light of serial homology. Nothing was seen where the main axis of the appendage should develop in abdominal segments. The primordia in A1 and A9 disappeared before larval hatching. Anal prolegs appeared separate from cerci, the main axes of appendages, which were formed temporarily in A11. The expression of decapentaplegic, which reflects the primary determination of appendages, was detected in the lateral juxtaposition with the prolegs. Distal-less was expressed in the main axes of appendages, protruding endites and the cerci, but not in prolegs and anal prolegs or the gnathal endites which do not protrude. These findings suggest a possibility that the abdominal and anal prolegs of A. rosae are outgrowths of ventral plates which derived from coxopodal elements, but not main axes of appendages.     

Characterization of Endoplasmic Reticulum-Localized UDP-d-Galactose: Hydroxyproline O-Galactosyltransferase Using Synthetic Peptide Substrates in Arabidopsis

We characterized peptidyl hydroxyproline (Hyp) O-galactosyltransferase (HGT), which is the initial enzyme in the arabinogalactan biosynthetic pathway. An in vitro assay of HGT activity was established using chemically synthesized fluorescent peptides as acceptor substrates and extracts from Arabidopsis (Arabidopsis thaliana) T87 cells as a source of crude enzyme. The galactose residue transferred to the peptide could be detected by high-performance liquid chromatography and matrix-assisted laser desorption-ionization time-of-flight mass spectrometry analyses. HGT required a divalent cation of manganese for maximal activity and consumed UDP-d-galactose as a sugar donor. HGT exhibited an optimal pH range of pH 7.0 to 8.0 and an optimal temperature of 35°C. The favorable substrates for the activity seemed to be peptides containing two alternating imino acid residues including at least one acceptor Hyp residue, although a peptide with single Hyp residue without any other imino acids also functioned as a substrate. The results of sucrose density gradient centrifugation revealed that the cellular localization of HGT activity is identical to those of endoplasmic reticulum markers such as Sec61 and Bip, indicating that HGT is predominantly localized to the endoplasmic reticulum. To our knowledge, this is the first characterization of HGT, and the data provide evidence that arabinogalactan biosynthesis occurs in the protein transport pathway.O-glycosylation is the addition of a sugar to hydroxy amino acids such as Thr, Ser, Hyp, Hyl, or Tyr (Lehle et al., 2006). This type of protein modification occurs in many organisms to modify a large variety of proteins. Several types of sugars can be linked to proteins via O-glycosylation, including Man, N-acetylgalactosamine, Glc, Xyl, N-acetylglucosamine, Fuc, Gal, and arabinofuranose (Araf). In addition, elongation of the added sugar residues yields a large variety of oligo- and polysaccharide extensions on the substrate proteins. These modifications are known to play important roles in various phenomena, including pathways required to maintain biological systems and basic cellular functions.Structural analysis of oligo- and polysaccharides in plant cell walls has revealed the presence of three types of O-linked structures, Gal-O-Hyp, Araf-O-Hyp, and Gal-O-Ser (Kieliszewski and Shpak, 2001; Seifert and Roberts, 2007). A part of these three structures has been found on proteins in the super family that includes arabinogalactan protein (AGP) and extensin, which are localized to the cell surface. AGPs contain O-linked arabinogalactan oligo- or polysaccharides attached to Hyp residues (Gal-O-Hyp). It is known that arabinogalactan polysaccharides mainly consist of β-1,3 linkages of Gal polymers (Seifert and Roberts, 2007). Extensin contains short arabino-oligosaccharide chains attached to Hyp residues (Araf-O-Hyp) and single Gal residues linked to Ser residues (Gal-O-Ser). It has been suggested that these O-linked structures play an important role in many stages of growth and development in plants, including signaling, embryogenesis, and programmed cell death (Knox, 2006; Seifert and Roberts, 2007). However, our understanding of the biosynthesis of these O-linked structures is limited at present.Shpak et al. described a novel strategy to elucidate O-glycosylation of AGPs via introduction of synthetic genes encoding a protein substrate of glycosyltransferases into plant cells (Shpak et al., 1999; Estevez et al., 2006). This strategy provided good evidence for the substrate specificities of Hyp O-galactosyltransferase (HGT). Hyp galactosylation occurs on clustered noncontiguous Hyp residues such as Xaa-Hyp-Xaa-Hyp repeats of AGPs (where Xaa is any amino acid except Hyp; Tan et al., 2003). However, the arabinogalactosylation site is not limited to clustered noncontiguous Hyp residues, as isolated Hyp residues with appropriate surrounding sequences can be modified with arabinogalactan (Matsuoka et al., 1995; Shimizu et al., 2005). Therefore, the mechanism of glycosylation to Hyp residues seems complex in plants, while we have little information about the glycosyltransferase(s) involved in arabinogalactan biosynthesis. To examine the enzymatic properties and to identify genes involved in arabinogalactan biosynthesis, we first attempted to establish an in vitro assay for HGT activity, which catalyzes the initial step in arabinogalactan biosynthesis in plants.Here, we report a novel assay for HGT activity based on the use of endoplasmic reticulum (ER)-enriched cell lysates extracted from Arabidopsis (Arabidopsis thaliana) T87 cells as a source of the enzyme and chemically synthesized fluorescent peptides as enzyme substrates. The method enabled us to characterize the enzymatic properties of HGT and to determine the localization of HGT in Arabidopsis cells. Properties of the enzyme and the usefulness of our assay for various studies are discussed.     

Tissue‐dependent induction of apoptosis by matrix metalloproteinase stromelysin‐3 during amphibian metamorphosis

Matrix metalloproteinases (MMPs) are a superfamily of Zn²⁺‐dependent proteases that are capable of cleaving the proteinaceous component of the extracellular matrix (ECM). The ECM is a critical medium for cell–cell interactions and can also directly signal cells through cell surface ECM receptors, such as integrins. In addition, many growth factors and signaling molecules are stored in the ECM. Thus, ECM remodeling and/or degradation by MMPs are expected to affect cell fate and behavior during many developmental and pathological processes. Numerous studies have shown that the expression of MMP mRNAs and proteins associates tightly with diverse developmental and pathological processes, such as tumor metastasis and mammary gland involution. In vivo evidence to support the roles of MMPs in these processes has been much harder to get. Here, we will review some of our studies on MMP11, or stromelysin‐3, during the thyroid hormone‐dependent amphibian metamorphosis, a process that resembles the so‐called postembryonic development in mammals (from a few months before to several months after birth in humans when organ growth and maturation take place). Our investigations demonstrate that stromelysin‐3 controls apoptosis in different tissues via at least two distinct mechanisms. Birth Defects Research (Part C) 90:55–66, 2010. © 2010 Wiley‐Liss, Inc.