Ozgur Cakici

Plasmalogen phospholipids protect internodal myelin from oxidative damage

Reactive oxygen species (ROS) are implicated in a range of degenerative conditions, including aging, neurodegenerative diseases, and neurological disorders. Myelin is a lipid-rich multilamellar sheath that facilitates rapid nerve conduction in vertebrates. Given the high energetic demands and low antioxidant capacity of the cells that elaborate the sheaths, myelin is considered intrinsically vulnerable to oxidative damage, raising the question whether additional mechanisms prevent structural damage. We characterized the structural and biochemical basis of ROS-mediated myelin damage in murine tissues from both central nervous system (CNS) and peripheral nervous system (PNS). To determine whether ROS can cause structural damage to the internodal myelin, whole sciatic and optic nerves were incubated ex vivo with a hydroxyl radical-generating system consisting of copper (Cu), hydrogen peroxide (HP), and ortho-phenanthroline (OP). Quantitative assessment of unfixed tissue by X-ray diffraction revealed irreversible compaction of myelin membrane stacking in both sciatic and optic nerves. Incubation in the presence of the hydroxyl radical scavenger sodium formate prevented this damage, implicating hydroxyl radical species. Myelin membranes are particularly enriched in plasmalogens, a class of ether-linked phospholipids proposed to have antioxidant properties. Myelin in sciatic nerve from plasmalogen-deficient (Pex7 knockout) mice was significantly more vulnerable to Cu/OP/HP-mediated ROS-induced compaction than myelin from WT mice. Our results directly support the role of plasmalogens as endogenous antioxidants providing a defense that protects ROS-vulnerable myelin.

A yeast-based chemical screen identifies a PDE inhibitor that elevates steroidogenesis in mouse Leydig cells via PDE8 and PDE4 inhibition.

A cell-based high-throughput screen (HTS) was developed to detect phosphodiesterase 8 (PDE8) and PDE4/8 combination inhibitors. By replacing the Schizosaccharomyces pombe PDE gene with the murine PDE8A1 gene in strains lacking adenylyl cyclase, we generated strains whose protein kinase A (PKA)-stimulated growth in 5-fluoro orotic acid (5FOA) medium reflects PDE8 activity. From our previously-identified PDE4 and PDE7 inhibitors, we identified a PDE4/8 inhibitor that allowed us to optimize screening conditions. Of 222,711 compounds screened, ∼0.2% displayed composite Z scores of >20. Additional yeast-based assays using the most effective 367 compounds identified 30 candidates for further characterization. Among these, compound BC8-15 displayed the lowest IC50 value for both PDE4 and PDE8 inhibition in in vitro enzyme assays. This compound also displays significant activity against PDE10A and PDE11A. BC8-15 elevates steroidogenesis in mouse Leydig cells as a single pharmacological agent. Assays using BC8-15 and two structural derivatives support a model in which PDE8 is a primary regulator of testosterone production by Leydig cells, with an additional role for PDE4 in this process. BC8-15, BC8-15A, and BC8-15C, which are commercially available compounds, display distinct patterns of activity against PDE4, PDE8, PDE10A, and PDE11A, representing a chemical toolkit that could be used to examine the biological roles of these enzymes in cell culture systems.

Crystal Structures of NodS N-Methyltransferase from Bradyrhizobium japonicum in Ligand-Free Form and as SAH Complex.

NodS is an S-adenosyl-L-methionine (SAM)-dependent N-methyltransferase that is involved in the biosynthesis of Nod factor (NF) in rhizobia, which are bacterial symbionts of legume plants. NF is a modified chitooligosaccharide (COS) signal molecule that is recognized by the legume host, where it initiates symbiotic processes leading to atmospheric nitrogen fixation. We report the crystal structure of recombinant NodS protein from Bradyrhizobium japonicum, which infects lupine and serradella legumes. Two crystal forms--ligand-free NodS and NodS in complex with S-adenosyl-L-homocysteine, which is a by-product of the methylation reaction--were obtained, and their structures were refined to resolutions of 2.43 Å and 1.85 Å, respectively. Although the overall fold (consisting of a seven-stranded β-sheet flanked by layers of helices) is similar to those of other SAM-dependent methyltransferases, NodS has specific features reflecting the unique character of its oligosaccharide substrate. In particular, the N-terminal helix and its connecting loop get ordered upon SAM binding, thereby closing the methyl donor cavity and shaping a long surface canyon that is clearly the binding site for the acceptor molecule. Comparison of the two structural forms of NodS suggests that there are also other conformational changes taking place upon the binding of the donor substrate. As an enzyme that methylates a COS substrate, NodS is the first example among all SAM-dependent methyltransferases to have its three-dimensional structure elucidated. Gaining insight about how NodS binds its donor and acceptor substrates helps to better understand the mechanism of NodS activity and the basis of its functional difference in various rhizobia.

Evaluation of ocular findings in patients with vitiligo.

In this study, we aimed to investigate ocular manifestations in patients with vitiligo. Sixty‐one patients with vitiligo were included in the study. From the patients who referred for examination to the dermatology and ophthalmology clinic, 57 patients without any systemic disease were taken as the control group. In both groups, otorefractometry, keratometry, visual acuity test, intraocular pressure measurement, anterior segment, and fundus examinations of the eye with slit lamp, Schirmer test, and perimetry were carried out. The mean age was 24.54 ± 11.90 years and 23.03 ± 8.72 years in the patients and control group, respectively. The mean Schirmer test results were as follows: 16.74 ± 9.11 mm and 17.64 ± 9.41 mm for the right and left eyes of the patients, and 21.96 ± 12.51 mm and 23.42 ± 12.51 mm for the right and left eyes of controls, respectively. Of the patients, 36 eyes showed lenticular findings. However, only 12 eyes of the controls have some lenticular findings. Twenty‐nine eyes in the vitiligo group and four in the controls showed some fundus findings. When the two groups were compared with each other, there was a statistically significant difference between them in terms of Schirmer test results, lens, and fundus findings (P < 0.05 for all). However, there was no significant difference in terms of age, gender, visual acuity, refraction, keratometry, intraocular pressure, perimetry, and corneal findings (P > 0.05 for all). Patients with vitiligo may have more lenticular and retinal findings than normal. They can be more prone to dry eye syndrome as well.

Arcuate keratotomy on post-keratoplasty astigmatism is unpredictable and frequently needs repeat procedures to increase its success rate.

Purpose To evaluate the effectiveness and predictability of arcuate keratotomy (AK) for post-keratoplasty astigmatism and to present the complications and rate of repeat procedures. Methods Sixteen eyes from 14 patients were included. Paired 70–80° arc length AKs centred on the steep axis were carried out 0.5 mm within the graft–host junction. The depth of the AKs was set at approximately 80–90% of the depth of the cornea, based on a topographic pachymeter at the incision location. The outcome measures included preoperative and postoperative topographic astigmatism, uncorrected and corrected visual acuity, surgical complications and repeat procedures. Results In 12 of the 16 eyes (75%), at least one additional surgical procedure was required to obtain the desired result: suturing for overcorrection or wound gape in six eyes (38%), lengthening of the incisions for undercorrection in four eyes (25%) and additional AKs for marked astigmatic axis displacement in three eyes (19%). The mean preoperative astigmatism was 10.45±3.82 dioptres (D); the postoperative astigmatism at the last visit was 2.99±1.14 D (in a mean follow-up of 17.6±5.55 months). The efficacy index was 0.83 and the safety index was 1.68. Conclusions In treatment of post-keratoplasty astigmatism, AK does not have a good predictability. Additional procedures such as lengthening of the AK incisions for undercorrection or using compression sutures for overcorrection with significantly gaping wounds are frequently required to improve the final outcome.

Periorbital discoid lupus: a rare localization in a patient with systemic lupus erythematosus

A 40-year-old female patient with a 5-year history of systemic lupus erythematosus was referred to our policlinic with complaints of erythema, atrophy, and telangiectasia on the upper eyelids for 8 months. No associated mucocutaneous lesion was present. Biopsy taken by our ophthalmology department revealed discoid lupus erythematosus. Topical tacrolimus was augmented to the systemic therapeutic regimen of the patient, which consisted of continuous antimalarial treatment and intermittent corticosteroid drugs. We observed no remission in spite of the 6-month supervised therapy. Periorbital discoid lupus erythematosus is very unusual and should be considered in the differential diagnosis of erythematous lesions of the periorbital area..

Sutureless scleral fixation of intraocular lenses

Dear Editor, In their article, Abbey et al. presented two sutureless scleral fixated intraocular lens methods they called BSutureless scleral fixation of intraocular lenses: outcomes of two approaches. The 2014 Yasuo Tano Memorial Lecture^ [1]. The authors stated that the trocar-assisted transconjunctival sutureless scleral fixation method had been described by Prasad [2]. In fact, we first described that method about 1 year earlier than Prasad [3]. In this technique, we enter into the eye with 25-gauge vitrectomy trocars without any opening in conjunctiva, performing two 3-mm-long scleral tunnels parallel to the limbus 180 degrees apart from each other, at an angle of 10 degrees to the scleral surface, and we leave the trocars there. Then we perform vitrectomy if necessary, and we grip and take the haptics out of the eye with retinal forceps through the trocars. The haptics are pushed to the end of the mouth of the tunnels, and then we put a 10/0 nylon suture encircling the haptics for security and take it out 1 week later [3]. The authors reported endophthalmitis in one case and hypotony in two cases [1]. In fact, we did not observe any cases of endophthalmitis or hypotony in our series. To us, the complications the authors encountered with their cannula technique are due to lack of the security suture at the end of the procedure. We think that, in our technique, by means of the 10/0 nylon security sutures in the scleral tunnels containing the haptics transconjunctivally, we ensure that the haptics are stabilized in the early postoperative period while we prevent possible leakage of the wound. So we have provided against endophthalmitis and hypotony. We take these sutures out after 1 week [3–10]. In addition, Abbey et al. observed haptic dislocation in three patients [1]. We did not see any haptic dislocation in our series. To us, the reason for that is that the authors performed scleral tunnels at an angle of 30 to 40 degrees. It may not be possible to successfully achieve 3-mm-long scleral tunnels with this level of angulation.Wemaintain the trocars at an angle of 10 degrees to the scleral surface while we create scleral tunnels, and thus wewere able to achieve tunnels about 3 mm long in which the haptics are placed. We also think that the security sutures help us to prevent this complication [3–10].

Manual tunnel incision cataract surgery with sandwich technique in eyes with microcornea

Manual tunnel incision cataract surgery with sandwich technique in eyes with microcornea The consultation section in the February 2014 issue involved a 41-year-old man with bilateral microcornea, inferonasal uveal coloboma, and mature cataract with modest inferior subluxation. 1 There is an erratum in the fourth paragraph of the case presentation. The text says “there is moderate pseudophakodonesis...” in this cataractous eye. However, the term should be phacodonesis since the eyewas cataractous, not pseudophakic. On therapeutic options, we generally agree with all the consultants as well as the case editor in terms of the preference of a scleral incision. Since the eye has a microcornea with a corneal diameter of 8.5 mm, any corneal incision will result in much more endothelial cell damage than a scleral, more peripheral incision and will compromise the corneal transparency. We would prefer a scleral tunnel incision in the case, as did the consultants. In addition, we would prefer the sandwich technique described by Fry. We began using manual

Manual tunnel incision cataract surgery with sandwich technique may be a rationale alternative for mature cataracts.

Dear Editor, We read with interest the article of Yang et al. appearing on April issue of Indian J Ophthalmology.[1] As the surgeons performing manual small incision cataract surgery since 1996,[2,3,4,5,6] we would like to note couple of points about Yang et al.s paper: There is an erratum on the results section of abstract. It has been written that “self-sealing wound was achieved in 112 eyes (98.2%).” However, the study had consisted 112 eyes. The correct number should be “110 eyes (98.2%).” This technique necessitates an incision at the location of 135° for right-handed surgeon and of 45° for left-handed surgeon. Hence, it will not be easy to perform it in the surgery of left eyes for right-handed surgeon and in the surgery of right eyes for left-handed surgeon, especially in the eyes with prominent eyebrows and big nose. It is not easy to arrange the incision according to steep-axis in Yang et al.s technique. Hence, the surgeon will not correct the preexisting astigmatism in some cases. Indeed, the authors has given a bigger mean value of postoperative astigmatisms (1.5 D) than of preoperative astigmatism (0.71 D). The authors implanted a polymethylmethacrylate (PMMA) intraocular lens (IOL). While PMMA IOLs are cheap IOLs, they have significant spherical aberration. Today, there are cheap foldable IOLs that do not have such significant spherical aberration and can be use for this type of surgery. Yang et al. perform a large, 7 mm capsulorrhexis. This can increases likelihood of the decentralization of the IOL and probably increased posterior capsule opacification rate since the capsular edges do not overlap the optic edge circumferentially. Due to this, we prefer a 6.0 mm diameter circular capsulorrhexis and an IOL of 6 mm optic diameter. We do not encounter with any difficulty to prolapse the nucleus into the anterior chamber with 6 mm capsulorrhexis. We have been preserving the manual small incision cataract surgery in hard, brunescent cataract cases and in the cataract cases of vitrectomized eyes in which we consider that phacoemulsification may be risky.[5,6] We think that sandwich technique presents some advantages in mature cataract cases. In this method, the nucleus firmly grasped between two instruments, irrigating vectis and spatula. So an incision length of a diameter of the nucleus or 1 mm more is enough to be able to extract the nucleus out to the eye. Furthermore, we consider that removing the nucleus via sandwiching it firmly between two instruments prevents the corneal endothelium more than extracting it via just only vectis or exerting a pressure on the scleral wound lip, since the spatula in front of the nucleus would ensure to stay it away from the endothelium. In conclusion, We congratulate Yang et al. for this novel and interesting technique, which is a valuable contribution, especially for the surgeons who do not have enough instrumental possibilities to carry out phacoemulsification in undeveloped areas.

Cloning, expression, purification, crystallization and preliminary X-ray analysis of NodS N-methyltransferase from Bradyrhizobium japonicum WM9.

The Nod factor (NF) is a rhizobial signal molecule that is involved in recognition of a legume host and the formation of root and stem nodules. Some unique enzymes are involved in the biosynthesis of NF, which is a variously but specifically substituted lipochitooligosaccharide. One of these enzymes is NodS, an N-methyltransferase that methylates end-deacetylated chitooligosaccharide substrates. In the methylation reaction, NodS uses S-adenosyl-L-methionine (SAM) as a methyl donor. To date, no structural information is available about NodS from any rhizobium. X-ray crystallographic studies of the NodS protein from Bradyrhizobium japonicum WM9, which infects the legumes lupin and serradella, have been undertaken. The nodS gene was cloned and the recombinant protein was expressed in Escherichia coli cells using natural amino acids and as an SeMet derivative. NodS without ligands was crystallized in the presence of PEG 3350 and MgCl(2). The protein was also crystallized in complex with S-adenosyl-L-homocysteine (SAH) in the presence of PEG 8000 and MgCl(2). SAH is produced from SAM as a byproduct of the methylation reaction. The crystals of apo NodS are tetragonal and diffracted X-rays to 2.42 A resolution. The NodS-SAH complex crystallizes in an orthorhombic space group and the crystals diffracted X-rays to 1.85 A resolution.