Displacement of Submacular Hemorrhage Using Subretinal Cocktail Injection versus Pneumatic Displacement: A Real-World Comparative Study.

INTRODUCTION: The objective of this study was to compare the outcome of submacular hemorrhage (SMH) displacement using pneumatic displacement with intravitreal expansile gas versus pars plana vitrectomy (PPV) with subretinal injection of tissue plasminogen activator (tPA), anti-vascular endothelial growth factor (VEGF) agent, and air as primary surgery. METHODS: Retrospective interventional case series of 63 patients who underwent surgical displacement of SMH secondary to neovascular age-related macular degeneration (nAMD) or polypoidal choroidal vasculopathy (PCV) from May 1, 2015, to October 31, 2022. Medical records were reviewed for diagnosis, logMAR visual acuity (VA), central subfield thickness (CST), and postoperative displacement rates and complications up to 12 months after operation. RESULTS: The diagnosis was nAMD in 24 (38.1%) and PCV in 39 (61.9%) eyes. There were 40 (63.5%) eyes in the pneumatic displacement group (38 received C3F8, 2 received SF6) and 23 (36.5%) eyes in the subretinal cocktail injection. Mean baseline VA was 1.46 and 1.62, respectively (p = 0.404). The subretinal injection group had more extensive SMH (p = 0.005), thicker CST (1,006.6 µm vs. 780.2 µm, p = 0.012), and longer interval between symptom and operation (10.65 vs. 5.53 days, p < 0.001). The mean postoperative VA at 6 months was 0.67 and 0.91 (p = 0.180) for pneumatic displacement and subretinal injection groups, respectively, though VA was significantly better in the pneumatic group at 12-month visit (0.64 vs. 1.03, p = 0.040). At least 10 mean change in VA were >10 letters gain in both groups up to 12 months. Postoperative CST reduction was greater (625.1 µm vs. 326.5 µm, p = 0.008) and complete foveal displacement (87.0% vs. 37.5%), p < 0.001, odds ratio [OR] = 11.1) and displacement to arcade or beyond (52.5% vs. 17.5%, p = 0.009, OR = 5.15) were more frequent in the subretinal injection group. Two patients with failed pneumatic displacement were successfully treated with subretinal cocktail injection as a second operation. CONCLUSION: Surgical displacement of SMH leads to clinically meaningful improvement in VA. PPV with subretinal cocktail injection is more effective than pneumatic displacement in displacing SMH with similar safety profile despite longer interval before operation, higher CST, and more extensive SMH at baseline. Retinal surgeons could consider this novel technique in cases with thick and extensive SMH or as a rescue secondary operation in selected cases.

Cyclic hematopoiesis in a mixed-breed dog: case report and brief review.

An 8-wk-old, male, mixed-breed puppy was adopted from a rescue organization. From the time of adoption, the puppy suffered episodes of illness affecting various organ systems, which resolved with supportive therapy but relapsed once medical therapy was discontinued. Review of the hematologic data revealed cyclic fluctuations in circulating blood cells. Cyclicity was most prominent in neutrophils, with recurrent severe neutropenia. Neutropenic episodes lasted 5-6 d, with regular cycles of 11-14 d between nadir neutrophil counts. Genetic testing determined that the patient was homozygous mutant for the frameshift mutation in the adaptor protein complex 3 ß-subunit (AP3B1) gene, originally identified in gray collies with cyclic hematopoiesis (CH). Pedigree information was not available, but the patient's features were phenotypically distinct from those of collies. We describe here a case of the AP3B1 mutation in a mixed-breed dog that did not resemble a collie, undescribed previously, to our knowledge. Our findings indicate that the AP3B1 mutation and CH are present within the general canine population and are not restricted to collies.

Pathology in Practice.

In collaboration with the American College of Veterinary Pathologists.

Bronchial brush cytology, endobronchial biopsy, and SALSA immunohistochemistry in severe equine asthma.

Horses with severe equine asthma (SEA), also known as heaves and recurrent airway obstruction, have persistent neutrophilic inflammation of the lower airways. Cytologic evaluation of bronchoalveolar lavage (BAL) fluid is commonly used to confirm the clinical diagnosis of SEA. However, the utility of microscopic assessment of bronchial brushings, endobronchial biopsies, and immunohistochemical detection of disease-associated biomarkers for the diagnosis of SEA remain poorly characterized. Salivary scavenger and agglutinin (SALSA) has anti-inflammatory properties and downregulated gene expression in SEA; therefore, it was investigated as a tissue biomarker for airway and systemic inflammation. Six asthmatic and 6 non-asthmatic horses were exposed to an inhaled challenge. Before and after challenge, samples of BAL fluid, bronchial brushing, and endobronchial biopsy were collected. Location of SALSA in biopsies was determined, and immunohistochemical label intensity was computed using image analysis software. Serum amyloid A (SAA) was measured to assess systemic inflammation. After challenge, neutrophil proportions were significantly higher in asthmatic versus non-asthmatic horses in BAL fluid (least squares means, 95% confidence interval: 80.9%, 57.2% to 93.1%, vs 3.6%, 1.1% to 10.7%) and in brush cytology slides (39.5%, 7.7% to 83.6%, vs 0.2%, 0% to 2.3%), illustrating the potential of brush cytology as an alternate modality to BAL for assessing intraluminal inflammation. Bronchial histopathologic findings and intensity of SALSA immunolabeling in surface and glandular epithelium were similar in asthmatic and non-asthmatic horses, indicating limited changes in bronchial tissue from the inhaled challenge. Increases in SAA indicated systemic inflammation, but SALSA immunolabeling did not change significantly.

Synthetic biology: at the crossroads of genetic engineering and human therapeutics-a Keystone Symposia report.

Synthetic biology has the potential to transform cell- and gene-based therapies for a variety of diseases. Sophisticated tools are now available for both eukaryotic and prokaryotic cells to engineer cells to selectively achieve therapeutic effects in response to one or more disease-related signals, thus sparing healthy tissue from potentially cytotoxic effects. This report summarizes the Keystone eSymposium "Synthetic Biology: At the Crossroads of Genetic Engineering and Human Therapeutics," which took place on May 3 and 4, 2021. Given that several therapies engineered using synthetic biology have entered clinical trials, there was a clear need for a synthetic biology symposium that emphasizes the therapeutic applications of synthetic biology as opposed to the technical aspects. Presenters discussed the use of synthetic biology to improve T cell, gene, and viral therapies, to engineer probiotics, and to expand upon existing modalities and functions of cell-based therapies.

Diversifying the structure of zinc finger nucleases for high-precision genome editing.

Genome editing for therapeutic applications often requires cleavage within a narrow sequence window. Here, to enable such high-precision targeting with zinc-finger nucleases (ZFNs), we have developed an expanded set of architectures that collectively increase the configurational options available for design by a factor of 64. These new architectures feature the functional attachment of the FokI cleavage domain to the amino terminus of one or both zinc-finger proteins (ZFPs) in the ZFN dimer, as well as the option to skip bases between the target triplets of otherwise adjacent fingers in each zinc-finger array. Using our new architectures, we demonstrate targeting of an arbitrarily chosen 28 bp genomic locus at a density that approaches 1.0 (i.e., efficient ZFNs available for targeting almost every base step). We show that these new architectures may be used for targeting three loci of therapeutic significance with a high degree of precision, efficiency, and specificity.

First report of Cryptosporidium parvum in a dromedary camel calf from Western Australia.

Cryptosporidium is an important enteric parasite that can contribute large numbers of infectious oocysts to drinking water catchments. As a result of its resistance to disinfectants including chlorine, it has been responsible for numerous waterborne outbreaks of gastroenteritis. Wildlife and livestock play an important role in the transmission of Cryptosporidium in the environment. Studies conducted outside Australia have indicated that camels may also play a role in the transmission of zoonotic species of Cryptosporidium. Despite Australia being home to the world's largest camel herd, nothing is known about the prevalence and species of Cryptosporidium infecting camels in this country. In the present study, C. parvum was identified by PCR amplification and sequencing of a formalin-fixed intestinal tissue specimen from a one-week old dromedary camel (Camelus dromedarius). Subtyping analysis at the glycoprotein 60 (gp60) locus identified C. parvum subtype IIaA17G2R1, which is a common zoonotic subtype reported in humans and animals worldwide. Histopathological findings also confirmed the presence of large numbers of variably-sized (1-3 µm in diameter) circular basophilic protozoa - consistent with Cryptosporidium spp.- adherent to the mucosal surface and occasionally free within the lumen. Further analysis of the prevalence and species of Cryptosporidium in camel populations across Australia are essential to better understand their potential for contamination of drinking water catchments.

Risk of recurrence of retinopathy of prematurity after initial intravitreal ranibizumab therapy.

We report our experience with the use of intravitreal ranibizumab for the treatment of retinopathy of prematurity (ROP). A retrospective review was performed on 138 consecutive infants screened at a single centre over 18 months. Intravitreal ranibizumab was offered in selected cases requiring treatment, such as aggressive posterior ROP or poor mydriasis. 2 eyes of 1 infant received intravitreal ranibizumab alone and 8 eyes of 5 infants received combined intravitreal ranibizumab and laser therapy. 3 out of 8 eyes treated initially with intravitreal ranibizumab monotherapy had persistent disease requiring laser therapy, and 3 out of 5 eyes with initial regression suffered disease recurrence at a mean of 7.6 weeks post-injection. 2 eyes treated first with laser followed by intravitreal ranibizumab had disease regression without recurrence. Our cohort demonstrate a significant rate of persistent disease and recurrence in ROP eyes treated initially with intravitreal ranibizumab monotherapy, which is greater and earlier than that reported for intravitreal bevacizumab in the BEAT-ROP study. Intravitreal ranibizumab may be useful as an initial treatment in selected cases of ROP when laser therapy as first line is suboptimal. However, close monitoring is important and adjunctive laser therapy may subsequently be needed in a majority of cases.

The natural history of polypoidal choroidal vasculopathy: a multi-center series of untreated Asian patients.

PURPOSE: We aimed to evaluate the long-term natural history of polypoidal choroidal vasculopathy (PCV) in untreated patients. METHODS: This is a retrospective observational case series. Patients with symptomatic PCV who did not receive any treatment for at least 12 months were included from the records of three ophthalmic clinics in Asia. The medical records and imaging data were reviewed. Visual outcomes at month 12 and at last follow-up were analyzed. The influence of demographics and presenting features on visual outcome was analyzed. RESULTS: A total of 32 eyes (32 patients) were included in this analysis. The mean follow-up was 59.9 months (range, 18-119 months), the mean age was 65.7 years and 21 (65.6 %) patients were male. The mean presenting logMAR visual acuity was 0.79 (Standard deviation [SD] 0.49). The center of the fovea was involved by the PCV complex in 25 eyes (78.1 %). The mean greatest linear dimension (GLD) of the PCV complex was 2584 µm (SD 880). Twenty-three eyes (71.9 %) had a cluster-of-grapes configuration on indocyanine green angiography. Leakage of fluorescein angiography was present in 29 eyes (90.6 %). The mean logMAR vision deteriorated from 0.79 at baseline to 0.88 at month 12 (p = 0.11), and further to 1.14 (p = 0.003) at the last follow-up. The proportion of eyes that improved, remained unchanged and worsened was 21.9 %, 31.3 % and 46.9 %, respectively, at month 12; and 28.1 %, 9.4 % and 62.5 %, respectively, at last follow-up. The proportion of eyes with logMAR vision worse than 1.0 was 28.1 % at presentation, and increased to 31.3 % at month 12 and further to 53.1 % at last follow-up. Reasons for poor vision were due to retinal, subretinal or vitreous hemorrhage, and retinal pigment epithelium (RPE) atrophy and scarring. None of the presenting features were found to significantly influence visual outcome. CONCLUSIONS: Half of eyes presenting with symptomatic PCV had a relatively benign course without treatment and some even had vision improvement. However, in the remaining eyes, vision deteriorated significantly, mainly due to hemorrhage and scarring. There may be subtypes of PCV with divergent natural history.

Use of intravitreal dexamethasone implant in patients with cataract and macular edema undergoing phacoemulsification.

PURPOSE: To study the safety and efficacy of intravitreal dexamethasone implant in patients with cataract and macular edema undergoing phacoemulsification and intraocular lens (IOL) implantation. METHODS: Twenty-four eyes with macular edema secondary to diabetic retinopathy (diabetic macular edema [DME]) and retinal vein occlusion (RVO) were retrospectively reviewed. These eyes underwent phacoemulsification with IOL implantation and intravitreal dexamethasone implant 0.7 mg at the same setting between September 2012 and September 2013. Demographic data, best-corrected visual acuity (BCVA), central macular thickness (CMT), intraocular pressure (IOP), surgical time, and complications were recorded. RESULTS: Twelve eyes had DME and 12 eyes had RVO (10 central RVO and 2 branch RVO). Median baseline logMAR BCVA was 1.0 (Snellen 20/200) and mean baseline CMT was 530.2 ± 218.9 µm. Median follow-up duration was 13 months. At last follow-up, median visual acuity improved significantly to 0.523 (Snellen 20/66) (p = 0.003) and CMT decreased to 300.7 ± 78.1 µm (p = 0.000). Median surgical time was 23 minutes. There were no intraoperative complications. In 12 eyes, macular edema recurred, requiring further treatment, and median time to recurrences was 21 weeks. One eye had raised IOP after second dexamethasone implant for recurrent macular edema. No major complication such as vitreous hemorrhage, retinal detachment, or endophthalmitis occurred. CONCLUSIONS: Combined cataract surgery with intravitreal dexamethasone implant seems to be safe and effective in treating patients with cataract and macular edema in this small case series. A larger prospective study with longer follow-up is necessary to demonstrate the long-term benefit of this combined procedure.

A large retinal capillary hemangioma in the anterior retina treated with photodynamic therapy.

PURPOSE: To report a case of a large anterior retinal capillary hemangioma (RCH) treated successfully with photodynamic therapy (PDT). METHODS: Case report. RESULTS: A 12-mm-large RCH located in the anterior retina, with vision-threatening exudative complications that had not responded to cryotherapy and repeated laser photocoagulations, was treated with PDT using verteporfin. Exudation regressed and tumor growth arrested after treatment. CONCLUSION: PDT can be delivered effectively to a lesion in the anterior retina and should be included as an option for treating anteriorly located RCH when conventional cryotherapy and laser photocoagulation fail.

Intravitreal ranibizumab with or without photodynamic therapy for the treatment of symptomatic polypoidal choroidal vasculopathy.

PURPOSE: To evaluate the efficacy of intravitreal ranibizumab with or without verteporfin photodynamic therapy (PDT) in the treatment of symptomatic polypoidal choroidal vasculopathy. METHODS: Twenty-three eyes of 23 patients received 3 monthly intravitreal ranibizumab injections with or without indocyanine green angiography-guided PDT at baseline. All patients had follow-up of ≥12 months. Visual and anatomical outcomes were compared between the two groups and a PDT monotherapy group. RESULTS: Seven eyes had ranibizumab monotherapy, 16 had combined ranibizumab injections and verteporfin PDT, and 12 had PDT monotherapy. At 3 months, the mean logarithm of minimal angle of resolution best-corrected visual acuity improved from 0.92 to 0.74 in the ranibizumab group (P = 0.18), from 0.70 to 0.59 in the combined group (P = 0.037), and from 0.74 to 0.57 in the PDT monotherapy group (P = 0.014). Complete regression of polypoidal lesions in indocyanine green angiography was found in 1 (14.3%) eye in the ranibizumab group, compared with 15 (93.8%) eyes in the combined group (P = 0.001). Additional PDT and ranibizumab injections in eyes with persistent polyps and fluorescein leakage resulted in regression of polyps in all eyes. At 12 months, no significant difference in logarithm of minimal angle of resolution best-corrected visual acuity and visual change was found between eyes initially treated with ranibizumab monotherapy, combined ranibizumab and PDT, or PDT monotherapy (P = 1.00 and P = 0.11, respectively). CONCLUSION: Intravitreal ranibizumab appeared to result in stabilization of vision in patients with symptomatic polypoidal choroidal vasculopathy. However, combined ranibizumab and PDT appeared to be more effective in causing complete regression of the polypoidal lesions in indocyanine green angiography compared with ranibizumab monotherapy.

Manganese superoxide dismutase and chemokine genes polymorphisms in chinese patients with anterior uveitis.

PURPOSE: To investigate the association of single-nucleotide polymorphisms (SNPs) in the manganese superoxide dismutase (MnSOD) and two chemokine genes (CCL2 and CCL5) in patients with anterior uveitis (AU). METHODS: Seventy-nine Chinese patients with acute AU were recruited, and genotyping of four SNPs including MnSOD 47, CCL2 -2518, CCL2 -2076, and CCL5 -403 alleles was performed with SNP genotyping assays. The genotype and allele frequencies were compared between patients with AU and 206 healthy control subjects. Analyses were also stratified according to the HLA-B27 status of the patients. RESULTS: There were significant increases in the frequency of the AA homozygosity in the MnSOD 47 SNP (P = 0.049) and in the CCL2 -2518G allele frequency and GG homozygosity in patients with AU compared with control subjects (P = 0.017 and P = 0.024, respectively). No significant association was found between AU with the CCL2 -2076 and CCL5 -403 SNPs. Subgroup analyses showed that the MnSOD 47A polymorphism was significantly associated with AU in HLA-B27-positive patients, but not in HLA-B27-negative patients, whereas the CCL2 -2518G polymorphism was significantly associated with AU in HLA-B27-negative patients, but not in HLA-B27-positive patients. CONCLUSIONS: The 47A polymorphism in the MnSOD gene and the -2518G polymorphism in the CCL2 gene are associated with the development of AU in HLA-B27-positive and -negative Chinese patients, respectively. Further studies to evaluate the interactions of the HLA-B27 status and these SNPs are warranted.

Repeatability and reproducibility of pachymetric mapping with Visante anterior segment-optical coherence tomography.

PURPOSE: To determine the repeatability and reproducibility of central and peripheral corneal pachymetry mapping with anterior segment-optical coherence tomography (AS-OCT). METHODS: An observational cross-sectional study involving two groups: 27 healthy eyes and 20 eyes with keratoconus. Each subject underwent scanning sessions with AS-OCT to determine intraobserver repeatability, interobserver reproducibility, and additionally for healthy eyes, intersession reproducibility for different regions of the cornea up to a 10-mm diameter. Main outcome measures were reproducibility and repeatability coefficients, intraclass correlation coefficients, and coefficients of variation of the average central (0-2 mm), pericentral (2-5 mm), transitional (5-7 mm), and peripheral (7-10 mm) corneal thicknesses generated by the Visante AS-OCT (Carl Zeiss Meditec, Inc., Dublin, CA) pachymetric mapping protocol. RESULTS: The coefficients of repeatability were less than 2% in healthy subjects and less than 3% in patients with keratoconus. The reproducibility coefficients were less than 2% and 4% in healthy subjects and patients with keratoconus, respectively. There was no significant difference between scans obtained by different observers or during different visits. The intraclass correlation coefficients were greater than 0.99 and 0.97 in healthy subjects and patients with keratoconus, respectively. CONCLUSIONS: With the pachymetric mapping protocol of Visante AS-OCT, these results suggest that central and peripheral corneal thickness measurements in healthy subjects and in eyes with keratoconus are repeatable and reproducible.

Retinal pigment epithelial tear following intravitreal ranibizumab injections for neovascular age-related macular degeneration.

BACKGROUND: To report the development of retinal pigment epithelial (RPE) tear after intravitreal injection of ranibizumab (Lucentis, Novartis, Basel, Switzerland). METHODS: Case report with presentation of the fundus photography, fluorescein angiography (FA) and optical coherence tomography (OCT) findings. RESULTS: A 70-year-old man received intravitreal injections of ranibizumab for the treatment of occult choroidal neovascularisation (CNV) with fibrovascular pigment epithelial detachment due to age-related macular degeneration. One day after the third intravitreal ranibizumab injection, fundus examination showed a RPE defect at the foveal region. FA and OCT confirmed the presence of RPE tear sparing the fovea. No further progression of the RPE tear was observed after withholding subsequent ranibizumab injection and his right eye visual acuity remained at 20/100 at 3 months from the last injection. CONCLUSIONS: As with other anti-vascular endothelial growth factor treatment for CNV, RPE tear might occur after intravitreal ranibizumab injection even after previous uneventful intravitreal injections.

Safety and efficacy of micro-incisional cataract surgery with bimanual phacoemulsification for white mature cataract.

BACKGROUND: To evaluate the safety and efficacy of micro-incisional cataract surgery (MICS) with bimanual phacoemulsification for the management of white mature cataract. METHODS: Twenty-five eyes in 25 patients with mature cataract were prospectively recruited to undergo MICS with bimanual phacoemulsification. Serial changes in best-corrected visual acuity (BCVA), central corneal thickness (CCT) and endothelial cell density (ECD) were compared using the Wilcoxon signed-rank test. RESULTS: MICS was successfully performed in 24 (96%) of the 25 eyes, with 1 eye requiring conversion to extracapsular cataract extraction due to radial tear during continuous curvilinear capsulorhexis. The median preoperative BCVA was hand movement. On day 1 postoperatively, the median BCVA improved to 0.6 (p < 0.001 compared with baseline). All patients had BCVA of 0.6 or better at 3 months after surgery (p < 0.001 compared with baseline). The mean increase in CCT at day 1 and week 1 postoperatively was 11.5 and 7.1%, respectively. The change in mean CCT was no longer significant at month 3 postoperatively (p = 0.82). The mean reduction in ECD at 3 months postoperatively was 7.8% (p = 0.037). None of the patients developed any postoperative complications. CONCLUSIONS: MICS with bimanual phacoemulsification appeared to be a promising alterative for the management of white mature cataract.

PEG conjugation moderately protects adeno-associated viral vectors against antibody neutralization.

AAV gene therapy vectors have significant clinical promise, but serum neutralization poses a challenge that must be overcome. We have examined the potential of conjugating the AAV surface with activated polyethylene glycol chains to protect the vector from neutralizing antibodies. Two key parameters were investigated: the polymer chain size and the PEG:lysine conjugation ratio. Transduction data revealed that the vector is fully infectious until a critical PEG conjugation reaction ratio was exceeded, and this critical level was found to vary with polymer chain size. At this key conjugation ratio, however, particles were moderately protected from serum neutralization, 2.3-fold over unmodified vector, demonstrating that there is a small window of PEGylation for which particles are still fully infective and benefit from antibody protection. TEM results and structural analysis indicate that the drop of infectivity as the PEG concentration is increased beyond the critical conjugation ratio may be due to a combination of steric interference with viral regions necessary for infection as well as reaction at important lysine residues. However, this first study analyzing the potential of PEG to protect AAV from serum neutralization shows that the approach has promise, which can be further enhanced if the locations of PEG attachment can be more finely controlled.