Diagnostic Accuracy of Perception Threshold Tracking in the Detection of Small Fiber Damage in Type 1 Diabetes.

AIM: An objective assessment of small nerve fibers is key to the early detection of diabetic peripheral neuropathy (DPN). This study investigates the diagnostic accuracy of a novel perception threshold tracking technique in detecting small nerve fiber damage. METHODS: Participants with type 1 diabetes (T1DM) without DPN (n = 20), with DPN (n = 20), with painful DPN (n = 20) and 20 healthy controls (HCs) underwent perception threshold tracking on the foot and corneal confocal microscopy. Diagnostic accuracy of perception threshold tracking compared to corneal confocal microscopy was analyzed using logistic regression. RESULTS: The rheobase, corneal nerve fiber density (CNFD), corneal nerve branch density (CNBD), and corneal nerve fiber length (CNFL) (all P < .001) differed between groups. The diagnostic accuracy of perception threshold tracking (rheobase) was excellent for identifying small nerve fiber damage, especially for CNFL with a sensitivity of 94%, specificity 94%, positive predictive value 97%, and negative predictive value 89%. There was a significant correlation between rheobase with CNFD, CNBD, CNFL, and Michigan Neuropathy Screening Instrument (all P < .001). CONCLUSION: Perception threshold tracking had a very high diagnostic agreement with corneal confocal microscopy for detecting small nerve fiber loss and may have clinical utility for assessing small nerve fiber damage and hence early DPN. CLINICAL TRIALS: NCT04078516.

The use of in vivo confocal microscopy in fungal keratitis - Progress and challenges.

Fungal keratitis (FK) is a serious and sight-threatening corneal infection with global reach. The need for prompt diagnosis is paramount, as a delay in initiation of treatment could lead to irreversible vision loss. Current "gold standard" diagnostic methods, namely corneal smear and culture, have limitations due to diagnostic insensitivity and their time-consuming nature. PCR is a newer, complementary method used in the diagnosis of fungal keratitis, whose results are also sample-dependent. In vivo confocal microscopy (IVCM) is a promising complementary diagnostic method of increasing importance as it allows non-invasive real-time direct visualization of potential fungal pathogens and manifesting infection directly in the patient's cornea. In numerous articles and case reports, FK diagnosis by IVCM has been evaluated, and different features, approaches, sensitivity/specificity, and limitations have been noted. Here, we provide an up-to-date, comprehensive review of the current literature and present the authors' combined recommendations for fungal identification in IVCM images, while also looking to the future of FK assessment by IVCM using artificial intelligence methods.

Validation of Catquest-9SF in Danish: developing a revised form of the Catquest-9SF - the Danish Catquest-7SF.

PURPOSE: The primary goal was to validate a Danish translated version of the Catquest-9SF by Rasch analysis. The secondary goal was to investigate whether preoperative Catquest-9SF scores, best-corrected visual acuity, comorbidity, gender, age or corneal astigmatism could predict improvements in subjective outcome. METHODS: In a prospective trial, 250 patients eligible for cataract surgery were included. Patients filled out the translated Catquest-9SF questionnaire before surgery and again 3 months after surgery. Both preoperative and postoperative questionnaires were included in the Rasch analysis. A multiple reverse stepwise regression model was used to investigate the correlation between preoperative measurements and subjective improvement. RESULTS: The preliminary Rasch analysis showed misfit of items 4 and 6. These items were removed, and the remaining seven items demonstrated a measurement precision of 2.78, a person reliability coefficient of 0.89, ordered response categories, infit of 0.69-1.22, outfit of 0.73-1.14, observed raw variance explained by measures of 70.4% and an eigenvalue of 1.7. Item 7 showed a mild DIF for gender (0.54 logits), and person mean Rasch score targeting was -1.69 logits. Preoperative Catquest score was the only parameter with a significant correlation to a gain in subjective outcome (p < 0.001). A preoperative Catquest-9SF score of 0.5 carried a 95% likelihood of an increase in subjective outcome. CONCLUSION: The Danish version of the Catquest-9SF fit the Rasch model. Only preoperative Catquest-9SF score was correlated to subjective improvement, and a cut-off value of 0.5 predicted an improvement in subjective outcome with 95% probability.

Fuchs' endothelial corneal dystrophy: a controlled prospective study on visual recovery after endothelial keratoplasty.

PURPOSE: To investigate the determining factors of vision and subjective outcome after Descemet's stripping automated endothelial keratoplasty (DSAEK) for Fuchs' endothelial dystrophy (FECD). METHODS: In a prospective study, 41 FECD patients who received DSAEK were compared to 40 cataract patients with normal corneas who received cataract surgery (control group). Subjects were recruited between March 2013 and July 2014. Observational procedures included the following: best-corrected visual acuity (BCVA), contrast sensitivity (CS), Catquest-9SF questionnaire, Scheimpflug tomography and anterior OCT. Examinations were carried out before surgery and at 3-, 6- and 12-month follow-up. Main outcome measures were associations between corneal optics and visual parameters, as well as subjective improvement (Catquest-9SF effect size) RESULTS: Best-corrected visual acuity (BCVA) negatively correlated with anterior higher-order aberrations (HOA's) (p < 0.001) and central corneal thickness (p = 0.001). Contrast sensitivity (CS) was negatively correlated with anterior HOA's (p = 0.002) and positively correlated with posterior densitometry (p = 0.008). Catquest-9SF effect size was 1.32 (CI: 1.0-1.6) in the control group, 1.84 (CI: 1.4-2.3) in FECD patients who received phacoemulsification and intra-ocular lens implantation in combination with DSAEK (n = 26) and 1.37 (CI: 0.6-2.1) in pseudophakic FECD patients who received DSAEK (n = 15). CONCLUSIONS: DSAEK surgery leads to considerable improvement in patient-reported outcome in FECD patients. There was a strong correlation between anterior HOA's and vision after DSAEK, suggesting that anterior corneal pathology constitutes the major limitation for visual recovery after DSAEK.

Evaluation of Endothelial Pump Function in Fuchs Endothelial Dystrophy Before and After Endothelial Keratoplasty.

PURPOSE: To evaluate the endothelial pump function in vivo after Descemet stripping automated endothelial keratoplasty (DSAEK). METHODS: In a prospective controlled trial, a group of 17 patients with Fuchs endothelial corneal dystrophy (FECD) eligible for DSAEK surgery and a group of 15 patients with cataract but with normal corneas eligible for cataract surgery (controls) were formed. A low oxygen-permeable contact lens was used to induce corneal edema. Changes in central corneal thickness were monitored as an indirect measure of endothelial cell pump function. Experiments were performed before surgery and repeated 12 months after surgery. RESULTS: Comparing the FECD and control groups before surgery, there was 24.8% (13.5-36.1) more edema in the FECD group after 2 hours (P < 0.001) and 19.9% (8.6-31.3) more edema in the FECD group after 3 hours (P < 0.001). In the FECD group, there was 15% (3.1-26.9) less edema after DSAEK than before surgery (P = 0.015) after 3 hours. Comparing the DSAEK-treated eyes with the control eyes 12 months after surgery; there was 12.8% (3.5-22.1) more edema in the DSAEK group after 2 hours (P = 0.007), but after 3 hours, the percentages of edema were similar (P = 0.11). CONCLUSIONS: Twelve months after DSAEK surgery, the grafted endothelium cleared the induced edema as fast as the control group, indicating a viable and near-normal endothelial pump function. However, significant differences in the deswelling patterns were detected, which may be caused by the added corneal stroma after DSAEK.

Fuchs Endothelial Corneal Dystrophy: A Systematic Immunofluorescence Study of Collagen Type VIII Suggests Heterogeneous Pathophysiology.

PURPOSE: The purpose of this study was to investigate the pathophysiologic heterogeneity of Fuchs endothelial corneal dystrophy (FECD). METHODS: We conducted a systematic immunofluorescence study on 39 Descemet membrane samples from FECD patients and compared these with 10 Descemet membrane samples from patients with pseudophakic bullous keratopathy (PBK) and 7 normal corneas. Samples were analyzed with immunofluorescence using antibodies to the α1-chain [collagen VIII α1-chain (COL8A1)] and α2-chain (COL8A2). Intensity of staining was assessed using a subjective grading scale from 0 to 3. The presence of specific staining patterns was noted. RESULTS: The overall distribution of COL8A1 staining intensity between groups was significantly different (P = 0.002). There was marked/intense staining in 85% (33/39) of the FECD samples, 40% (4/10) of the PBK samples (P = 0.034), and 29% (2/7) of normal samples (P = 0.004). The overall distribution of COL8A2 staining intensity was not significantly different between groups (P = 0.39). There was marked/intense staining in 33% (13/39) of the FECD samples, 10% (1/10) of PBK samples, and 14% (1/7) of the normal samples. There was substantial variation in staining intensity in the FECD group, a phenomenon that was especially pronounced for the COL8A2 antibody. CONCLUSIONS: We found increased staining for COL8A1, but not COL8A2 in FECD samples. Further, there was striking variation of staining intensity in FECD patients, indicating pathophysiological heterogeneity.

The Ultrastructures and Mechanical Properties of the Descement's Membrane in Fuchs Endothelial Corneal Dystrophy.

Fuchs endothelial corneal dystrophy (FECD), is the most common corneal endothelial dystrophy, and contributes up to 50% of all corneal transplantations performed in developed countries. FECD develops in Descemet's membrane (DM) and possibly alters the mechanical properties and internal structures in this basal lamina. In this work, the morphology and mechanical properties of FECD-DMs are studied by transmission electron microscopy (TEM) and quantitative dynamic atomic force microscopy (QD-AFM) at nano scale. Pathological wide-space collagens that are typical of FECD display different mechanical properties in that they are softer than the remaining tissue both for dehydrated- and fully hydrated samples. Additionally, the hydration level has major influence on the mechanical properties. These findings could help to further understand the structural changes in FECD, and possibly be useful for further characterization of the disease, the diagnosis and assessment or even pathologic analysis.

LASIK surgery of granular corneal dystrophy type 2 patients leads to accumulation and differential proteolytic processing of transforming growth factor beta-induced protein (TGFBIp).

More than 60 mutations in transforming growth factor beta-induced protein (TGFBIp) have been reported in humans causing a variety of phenotypic protein aggregates in the cornea, commonly termed corneal dystrophies. One mutation, generating an arginine to histidine amino acid substitution at position 124 in mature TGFBIp leads to granular corneal dystrophy type 2 (GCD2). Homozygous GCD2 cases develop massive protein accumulation early in life whereas heterozygous GCD2 cases become affected much later and generally with a much less severe outcome. However, if heterozygous GCD2 patients undergo laser-assisted in situ keratomileusis (LASIK) surgery protein accumulation is accelerated and they develop massive protein accumulations a few years after surgery. Here, we present the protein profile of aggregate-containing corneal tissue from GCD2 patients with a history of LASIK surgery using LC-MS/MS. Label-free quantification of corneal extracellular matrix proteins showed accumulation of TGFBIp. This was supported by 2DE and immunoblotting against TGFBIp that revealed the accumulation of full-length TGFBIp. In addition, a high molecular weight TGFBIp complex was more apparent in GCD2 patients after LASIK surgery, which may be important for the disease progression. Lastly, 2DE also revealed differential processing between GCD2 patients with a history of LASIK surgery when compared to healthy individuals.

Incidence and clinical characteristics of fungal keratitis in a Danish population from 2000 to 2013.

PURPOSE: Fungal keratitis is a severe sight-threatening condition. The aim of this study was to investigate the incidence and clinical characteristics of fungal keratitis patients living in a temperate climate. METHODS: By reviewing medical records from 2000 to July 2013, patients with fungal keratitis were identified. Risk factors, clinical signs and outcome were registered. RESULTS: Twenty-five patients were identified: 52% with Candida, 20% with Fusarium, 16% with Aspergillus and 12% with mixed filamentous fungi. A minimum incidence of fungal keratitis of 0.6 cases per million per year was estimated. Prior topical steroid treatment was commonly found in our cases (44%). Trauma including contact lens wear was associated with infection with filamentous fungi, whereas in patients with Candida infection, ocular surface disease was a prominent feature. Median time from onset of symptoms to diagnosis was 24 days. Only a few patients exhibited classical clinical features such as endothelial plaques (28%), satellite lesions (24%) and feathery edges (16%). The final visual outcome was poor with an average best-corrected logMAR of (mean, 95% CI) 0.70 (0.4-1.0). A total of 52% were treated with corneal transplantation. Patients with Candida infections had a significantly worse visual outcome. CONCLUSION: We found that patients with fungal keratitis had a poor visual outcome. However, knowledge of risk factors and clinical signs leading to early treatment can improve the prognosis.

Additive effect of PTH (1-34) and zoledronate in the prevention of disuse osteopenia in rats.

Immobilization is known to cause a rapid bone loss due to increased osteoclastic bone resorption and decreased osteoblastic bone formation. Zoledronate (Zln) is a potent anti-resorptive pharmaceutical, while intermittent PTH is a potent bone anabolic agent. The aim of the present study was to investigate whether PTH or Zln alone or in combination could prevent immobilization-induced osteopenia. Immobilization was achieved by injecting 4IU Botox (BTX) into the right hind limb musculature. Seventy-two 16-week-old female Wistar rats were randomized into 6 groups; baseline (Base), control (Ctrl), BTX, BTX+PTH, BTX+Zln, and BTX+PTH+Zln. PTH (1-34) (80µg/kg) was given 5days/week and Zln (100µg/kg) was given once at study start. The animals were killed after 4weeks of treatment. The bone properties were evaluated using DEXA, µCT, dynamic bone histomorphometry, and mechanical testing. BTX resulted in lower femoral trabecular bone volume fraction (BV/TV) (-25%, p<0.05), lower tibial trabecular bone formation rate (BFR/BS) (-29%, p<0.05), and lower bone strength (Fmax) at the distal femur (-19%, p<0.001) compared with Ctrl. BTX+PTH resulted in higher femoral BV/TV (+31%, p<0.05), higher tibial trabecular BFR/BS (+297%, p<0.05), and higher Fmax at the distal femur (+11%, p<0.05) compared with BTX. BTX+Zln resulted in higher femoral BV/TV (+36%, p<0.05), lower tibial trabecular BFR/BS (-93%, p<0.05), and higher Fmax at the distal femur (+10%, p<0.05) compared with BTX. BTX+PTH+Zln resulted in higher femoral BV/TV (+70%, p<0.001), higher tibial trabecular BFR/BS (+59%, p<0.05), and higher Fmax at the distal femur (+32%, p<0.001) compared with BTX. In conclusion, BTX-induced immobilization led to lower BV/TV, BFR/BS, and Fmax. In general, PTH or Zln alone prevented the BTX-induced osteopenia, whereas PTH and Zln given in combination not only prevented, but also increased BV/TV and BFR/BS, and maintained Fmax at the distal femoral metaphysis compared with Ctrl.

Inverse cutting of posterior lamellar corneal grafts by a femtosecond laser.

PURPOSE: Posterior lamellar grafting of the cornea has become the preferred technique for treatment of corneal endothelial dysfunction. Posterior lamellar grafts are usually cut by a micro-keratome or a femto-second laser after the epithelial side of the donor cornea has been applanated. This approach often results in variable central graft thickness in different grafts and an increase in graft thickness towards the periphery in every graft. The purpose of this study was to evaluate if posterior lamellar grafts can be prepared from the endothelial side by a femto-second laser, resulting in reproducible, thin grafts of even thickness. METHODS: A CZM 500 kHz Visumax femto-second laser was used. Organ cultured donor grafts were mounted in an artifical anterior chamber with the endothelial side up and out. Posterior grafts of 7.8 mm diameter and 130 micron thickness were prepared by femto-second laser cutting. A standard DSAEK procedure was performed in 10 patients with Fuchs endothelial dystrophy. Patients were followed-up regularly and evaluated by measurement of complications, visual acuity, corneal thickness (Pentacam HR), and endothelial cell density. RESULTS: Femto-laser cutting of grafts and surgery was uncomplicated. Rebubbling was necessary in 5 of 10 cases (normally only in 1 of 20 cases). All grafts were attached and cleared up during the first few weeks. After six months, the average visual acuity was 0.30 (range: 0.16 to 0.50), corneal thickness was 0.58 mm (range 0.51 to 0.63), and endothelial cell density was 1.570 per sq. mm (range: 1.400 to 2.000 cells per sq. mm). The grafts were of uniform thickness, but substantial interface haze was present in most grafts. CONCLUSIONS: Posterior lamellar corneal grafts can be prepared from the endothelial side using a femto-second laser. All grafts were clear after 6 months with satisfying endothelial cell counts. Poor visual acuity caused by interface scatter was observed in most patients. Femto-second laser cutting parameters needs to be optimised to enable smooth cutting in the posterior stroma.

Visual acuity and contrast sensitivity after posterior lamellar keratoplasty.

PURPOSE: It has previously been reported that patients who suffer from Fuchs' endothelial dystrophy (Fuchs' ED) have decreased contrast sensibility threshold. The removal of endothelial guttata by Descemet's Stripping Automated Endothelial Keratoplasty (DSAEK) surgery has also been demonstrated to decrease intraocular light scatter and improve contrast thresholds. The purpose of this study was to compare different visual qualities in patients that had undergone DSAEK surgery in one eye while having untreated Fuchs' dystrophy in the other eye. METHODS: Thirty-two eyes of 16 patients with bilateral Fuchs' ED who had DSAEK surgery performed in one eye were enrolled. Visual acuity at 100% contrast and contrast sensitivity (CS) as evaluated by a modified simulation of the Freiburg Acuity and Contrast Test, was measured in both eyes of each patient. RESULTS: Snellen visual acuity improved in treated eyes from 0.66 ± 0.35 (SD) before surgery to 0.38 ± 0.18 after surgery (log MAR units; p < 0.01). In eyes with untreated Fuchs' dystrophy, visual acuity was 0.46 ± 0.14. In a pairwise comparison, there was no difference in Snellen visual acuity between treated and nontreated eyes (p = 0.12). Contrast sensitivity was significantly better in DSAEK-treated eyes compared with untreated eyes. Mean log CS in DSAEK-treated eyes was 1.06 ± 0.25 compared with a mean log CS of 0.84 ± 0.16 in untreated eyes (p < 0.01). Fourteen of 16 patients (88%) reported that vision in their DSAEK-treated eye had the best visual quality. CONCLUSION: In this study, using a paired design, we demonstrate improved CS in DSAEK-operated eyes when compared with the other eye that had untreated Fuchs' ED despite the fact that no difference in visual quality was detected with standard Snellen visual acuity. Contrast sensitivity appears to be superior to normal visual acuity measurements in the evaluation of overall visual quality after DSAEK.

[Early stage of Acanthamoeba keratitis]. / Tidligt stadie af akantamøbekeratitis.

We present a case of early detected Acanthamoeba keratitis. The patient was a 31 year-old female with keratitis who was referred to our clinic. Upon arrival the patient presented with subepithelial corneal infiltrates and radiating neuritis. In vivo confocal microscopy showed Acanthamoeba cysts and treatment was initiated. Polymerase chain reaction analysis later confirmed the diagnosis and after three months symptoms had abated and vision was 1.0 (6/6). We confirm that an early diagnosis af Acanthamoeba keratitis is pivotal.

[New diagnostic update in Acanthamoeba keratitis]. / Nye diagnostiske tiltag ved akantamøbekeratitis.

Acanthamoeba keratitis is a rare but sight threatening condition. A major problem is that the disease is difficult to diagnose and often mistaken for herpes infection in its early stages. We present an update in diagnostics with the presentation of five recent cases of Acanthamoeba keratitis where the use of confocal microscopy and PCR based DNA analysis of epithelial scrapings played a pivotal role. An early diagnose is crucial for achieving a successful outcome.

Microbial biomass C, N and P in two arctic soils and responses to addition of NPK fertilizer and sugar: implications for plant nutrient uptake.

The soil microbial carbon (C), nitrogen (N) and phosphorus (P) pools were quantified in the organic horizon of soils from an arctic/alpine low-altitude heath and a high-altitude fellfield by the fumigation-extraction method before and after factorial addition of sugar, NPK fertilizer and benomyl, a fungicide. In unamended soil, microbial C, N and P made up 3.3-3.6%, 6.1-7.3% and 34.7% of the total soil C, N and P content, respectively. The inorganic extractable N pool was below 0.1% and the inorganic extractable P content slightly less than 1% of the total soil pool sizes. Benomyl addition in spring and summer did not affect microbial C or nutrient content analysed in the autumn. Sugar amendments increased microbial C by 15 and 37% in the two soils, respectively, but did not affect the microbial nutrient content, whereas inorganic N and P either declined significantly or tended to decline. The increased microbial C indicates that the microbial biomass also increased but without a proportional enhancement of N and P uptake. NPK addition did not affect the amount of microbial C but almost doubled the microbial N pool and more than doubled the P pool. A separate study has shown that CO2 evolution increased by more than 50% after sugar amendment and by about 30% after NPK and NK additions to one of the soils. Hence, the microbial biomass did not increase in response to NPK addition, but the microbes immobilized large amounts of the added nutrients and, judging by the increased CO2 evolution, their activity increased. We conclude: (1) that microbial biomass production in these soils is stimulated by labile carbon and that the microbial activity is stimulated by both labile C and by nutrients (N); (2) that the microbial biomass is a strong sink for nutrients and that the microbial community probably can withdraw substantial amounts of nutrients from the inorganic, plant-available pool, at least periodically; (3) that temporary declines in microbial populations are likely to release a flush of inorganic nutrients to the soil, particularly P of which the microbial biomass contained more than one third of the total soil pool; and (4) that the mobilization-immobilization cycles of nutrients coupled to the population dynamics of soil organisms can be a significant regulating factor for the nutrient supply to the primary producers, which are usually strongly nutrient-limited in arctic ecosystems.