Cattle Retina Research Articles

CattNIS: Novel identification system of cattle with retinal images based on feature matching method

Small and large farm animals are frequently identified using techniques like microchipping or tagging. However, these techniques may have a detrimental impact on animal welfare and may result in theft and fraud. Modern and sophisticated identification procedures using biometrics, including retina, face, and iris recognition, are available and have less of a detrimental effect on animal welfare. Because biometric markers are unchangeable and unaffected, it is feasible to stop situations of theft and fraud. This study uses retina images to identify animals based on each animal's different and specific retinal patterns. To the best of our knowledge, there isn't a publicly accessible dataset for images of animal retinas. The three main goals of this study are to: 1) produce a novel dataset of Cattle retinal images and make it publicly available; 2) create a successful image processing-based system for retinal identification and recognition; and 3) create a graphical user interface (GUI) that enables querying the database to ascertain whether a specific Cattle retina is present. To accomplish these goals, a dataset comprising 2430 images from the left and right eyes of 300 cattle was produced. After the dataset was created, the animal retinal images were segmented using image processing methods such as scaling, color transformations, image sharpening, contrast enhancements, noise filtering, and histogram equalization. By using the segmented images with the SURF, FAST, BRISK, and HARRIS techniques, distinctive retinal features were found. During the identification phase, the animal whose retinal characteristic matched the sought animal's retina the closest was recognized. Within the developed CattNIS system, the SURF approach has the highest accuracy rate of 92.25 percent. The results of this study show that the identification of cattle using retinal vascular patterns is highly successful. The availability of the collected cattle retinal images from this study is expected to significantly contribute to the advancement of future research in this field, both in terms of quantity and quality.

Neuroprotection in the complex treatment of primary glaucoma

Neuroprotection is a direction in therapy that prevents the loss of neurons due to external factors. In glaucoma, the use of both direct and indirect neuroprotection is pathogenetically justified, which, respectively, can have a direct protective effect on retinal ganglion cells, or affect various risk factors for the development of neuropathy. At present, Retinalamin — a complex of peptides isolated from cattle retinas — is used in ophthalmology for neuroprotective purposes. Although the clinical efficacy and safety of Retinalamin is well established, its exact mechanism of action remains unclear. To amend this, in vitro studies of its cytotoxicity and efficacy were carried out, as well as a comparison of its effectiveness in various ways of administration, and a retrospective analysis of its use.

Evaluation of therapeutic sensitivity of retinal ganglion cells to targeted peptide bioregulator in culture

To study how therapeutically sensitive retinal cell culture is to the peptide bioregulator isolated from cattle retina (Retinalamin) in models of glaucomatous optic neuropathy (GON). The cells were isolated from the retinae separated from newborn mice. Cell sheets were disaggregated and transformed into a suspension. Removal of the off-target cell population was done by adding antibodies to deplete the cells with CD48 marker, and magnetic microbeads that attach to them. Selection of ganglion cells and obtainment of its enriched fraction was done by immunomagnetic separation. To assess the toxicity of Retinalamin, a cytotoxic test was performed on the culture of skin fibroblasts with sequential dilution of the drug into concentrations of 5.0-0.009 mg/mL. The cells were seeded at 5000 per plate well and exposed to the drug for 24 hours. To study the excitotoxic damage, the first group of plate wells with retinal cells had solution of sodium glutamate added in concentration of 20 mM, Retinalamin was added into the second group of wells in concentration of 1.25 mg/mL; both substances were added into the third group of wells. The control group consisted of intact plate wells. The cells were exposed to substances for 24 hours. Cell vitality was then evaluated using colorimetry. Optic density was measured using an automatic photometer with detection wavelength of λ=490 nm. The cell culture achieved by immunomagnetic separation is mixed and consists of ganglion and glial cells. Retinalamin does not display significant cytotoxicity in any of the studied concentrations. The excitotoxic damage caused significant decrease of the amount of viable cells in the culture (9% of the control wells). The concomitant addition of glutamate and 1.25 mg/mL Retinalamin resulted in a 51.6% increase in the amount of viable cells. The intergroup differences were statistically significant by Student's t-test. Retinalamin is not cytotoxic. The concomitant addition of glutamate and Retinalamin reliably decreases the toxic action of glutamate on isolated retinal cells.

Molecular mechanisms of neuroretinoprotection in primary open-angle glaucoma

To evaluate the effectiveness of neuroprotective therapy in patients with primary open-angle glaucoma based on specific biochemical markers of neurotrophy and neurodegeneration. In the course of the study, lacrimal fluid (LF) of 23 patients (46 eyes) with stage I-II primary open angle glaucoma (POAG) aged 66.3±10.8 was examined. The control group consisted of healthy individuals (12 control LF samples). Local antihypertensive therapy was complimented with 10 intramuscular injections of retinalamin (GEROPHARM LLC, St. Petersburg) - a complex of peptides isolated from cattle retinas, administered 5 mg once a day. Enzyme immunoassay (ELISA) method was used to test LF for concentrations of brain-derived neurotrophic factor (BDNF) and neuron specific enolase (NSE) before and after the treatment. In healthy subjects, the content of BDNF was 0.83±0.06 ng/mL, NSE - 0.51±0.06 ng/mL. In patients with POAG, the indices were higher: BDNF was 1.39±0.85 ng/mL (p>0.05), NSE - 4.31±2.02 ng/mL (p<0.05). The patients with stage II-III POAG had a significant increase of BDNF concentration (1.37±0.41 ng/mL and 1.52±0.40 ng/mL respectively in stages II and III) indicating compensatory capabilities at these stages of the disease. The NSE marker was also high in these stages (4.16±2.44 ng/mL and 5.78±2.80 ng/mL, p<0.05), which indicates the degeneration of retinal ganglion cells. After Retinalamin® therapy, patients with stage II POAG had control group levels of marker proteins, while in stage III patients NSE content remained high. There is a compensatory increase of BDNF level and a pathological increase of NSE content in LF of POAG patients. The change in the content of neurotrophins in the LF was determined to depend on the stage of POAG. Thus, in stages II and III POAG there is a marked increase of NSE and a compensatory increase of BDNF. After the complex therapy including retinalamin in patients with stage II POAG, a decrease in initially high concentrations of BDNF and NSE in LF down to the control group values was noted. In stage III patients, signs of neurodegeneration persist in the form of high NSE values. Taking into account the positive dynamics of the neuronal marker content in LF in the complex therapy of POAG with Retinalamin®, timely neuroretinoprotection is recommended.

Measurement of Blood Vitamin A Level for Beef Cattle Using Color Images of the Pupil and Retina

Marbling score is an important index of beef quality. Vitamin A (VA) in blood inhibits differentiation of preadipose cells into adipose cells. High VA level lowers the marbling score of beef. Therefore, breeders are trying to optimize the blood VA level to improve marbling score (ca. 30 IU/dl). Monitoring the blood VA level is essential, however, a currently available method of blood VA level is blood sampling which is expensive, invasive, and time-consuming. Therefore, an alternative method is desirable. In the previous study, there was a linear relationship between the blood VA level of cattle and the light intensity at 470nm which was reflected from cattle eye. The objective of this study was to investigate the relationship between the blood VA level and the color of cattle eye, especially the pupil color. The final goal of this project is to develop a non-invasive monitoring system of VA level in cattle blood using image processing technique. The image of cattle eye was captured using a CCD camera and analyzed with WinRoof software. The relationships between the blood VA level and the mean RGB value or chromaticity value, “rgb„, or HIS value of pupil were examined for twenty-five cattle with different blood VA level which were ranged between 15 IU/dl and 120 IU/dl. The image of cattle retina was also captured and analyzed in the same way. The blood VA level of test cattle was ranged between 15 IU/dl to 105 IU/dl. Comparing the pupil data with the retina data, Intensity value (I) showed a similar tendency for both pupil and retina images.

Retinal Function and Morphology Are Altered in Cattle Infected with the Prion Disease Transmissible Mink Encephalopathy

Transmissible spongiform encephalopathies (TSEs) are a group of diseases that result in progressive and invariably fatal neurologic disease in both animals and humans. TSEs are characterized by the accumulation of an abnormal protease-resistant form of the prion protein in the central nervous system. Transmission of infectious TSEs is believed to occur via ingestion of prion protein-contaminated material. This material is also involved in the transmission of bovine spongiform encephalopathy ("mad cow disease") to humans, which resulted in the variant form of Creutzfeldt-Jakob disease. Abnormal prion protein has been reported in the retina of TSE-affected cattle, but despite these observations, the specific effect of abnormal prion protein on retinal morphology and function has not been assessed. The objective of this study was to identify and characterize potential functional and morphologic abnormalities in the retinas of cattle infected with a bovine-adapted isolate of transmissible mink encephalopathy. We used electroretinography and immunohistochemistry to examine retinas from 10 noninoculated and 5 transmissible mink encephalopathy-inoculated adult Holstein steers. Here we show altered retinal function, as evidenced by prolonged implicit time of the electroretinogram b-wave, in transmissible mink encephalopathy-infected cattle before the onset of clinical illness. We also demonstrate disruption of rod bipolar cell synaptic terminals, indicated by decreased immunoreactivity for the alpha isoform of protein kinase C and vesicular glutamate transporter 1, and activation of Müller glia, as evidenced by increased glial fibrillary acidic protein and glutamine synthetase expression, in the retinas of these cattle at the time of euthanasia due to clinical deterioration. This is the first study to identify both functional and morphologic alterations in the retinas of TSE-infected cattle. Our results support future efforts to focus on the retina for the development of new strategies for the diagnosis of TSEs.

Quantitative analysis of very long chain fatty acids of bovine retina

Background. Bovine retina is known to contain very long chain polyunsaturated fatty acids (VLCPUFA) of chain length >24 carbons and 4–6 double bonds. We carefully quantified the ELCPUFA of bovine retina using modern methods in beef cattle. Methods. Bovine retinas were obtained from a slaughterhouse in Nebraska. Retinas were carefully dissected and 50 were pooled for analysis. Samples were extracted, the phosphatidyl choline (PC) fraction purified, and fatty acid methyl esters prepared. Structures were analyzed by covalent adduct chemical ionization tandem mass spectrometry, and quantified by capillary gas chromatography. High precision isotope ratios of 22:6n-3 and 20:4 were determined. Results. About 1% of the PC were VLCPUFA. They were detected at chain lengths from C24 to C36 and up to 6 double bonds. n-6 LCPUFA were high, especially 20:4 and 22:5, compared to studies published in the 1980s, probably reflecting the high n-6 content of the corn diet. 32:4n-6 was anomalously predominant compared to similar chain lengths and degrees of unsaturation. The δ13C for 20:4n-6 was 10‰ greater than for 22:6n-3. Conclusions. These data form a basis for models of membrane phenomena such as protein-phospholipid interactions that may rely on PUFA and VLCPUFA for activity. They also suggest that the retinas of corn-fed cattle are particularly low in n-3 LCPUFA. Funding: NIH GM071534.

Phosphatidylinositol-4,5-bisphosphate phospholipase C in bovine rod outer segments

Preparations of rod outer segments from cattle retinas contained soluble and particulate phospholipase C activities which hydrolyzed phosphatidylinositol 4,5-bisphosphate (PIP2) and the other phosphoinositides. Ca2+ was required for PIP2 hydrolysis, but high (greater than 300 microM) concentrations were inhibitory. Mg2+ and spermine at low concentrations stimulated the particulate activity but inhibited the soluble. Mn2+ inhibited both. High (greater than 100 microM) concentrations of the nonhydrolyzable GTP analogue guanylyl beta,gamma-methylenediphosphonate inhibited PIP2 hydrolysis by both the soluble and particulate activities, but guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S), fluoride, and cholera and pertussis toxins were without effect. Overall phospholipase C activity in ROS was unaffected by light. Evidence was found for multiple forms of the enzyme, requiring isolation and separate characterization before ruling out regulation by light or G-protein.

LOW‐TEMPERATURE PHOTOACOUSTIC SPECTROSCOPY OF PORPHYROPSIN IN THE INTACT RETINA OF THE CREEK CHUB, Semotilus atromaculatus

Abstract— The photoacoustic spectra of porphyropsin in the creek chub (Semotilus atromaculatus) have been successfully measured on single retinae of small size (&lt; 0.5 cm in diameter) at low temperatures. The relative intensities of the spectra, corresponding to the thermal release, are strong enough to be comparable to those for the big cattle retina. The photoacoustic spectrum of porphyropsin at – 167°C exhibits a principal maximum at 540 nm with a small shoulder near 580 nm. A sequence of porphyropsin bleaching resulted from thermal reaction has been clearly observed above ‐90°C.

Cyclic GMP phosphodiesterase from cattle retina: Amino acid sequence of the γ-subunit and nucleotide sequence of the corresponding cDNA

The primary structure of the γ-subunit of cyclic GMP phosphodiesterase was determined by parallel analysis of the amino acid sequence of the protein and nucleotide sequence of the corresponding cDNA. The enzyme γ-subunit contains 87 amino acid residues, its N-terminal amino group being acetylated.

Depth-resolved chromophore analysis of bovine retina and pigment epithelium by photoacoustic spectroscopy

The cattle retina and retinal pigment epithelium are examined by means of photoacoustic spectroscopy under conditions where structural integrity is maintained. Depth-related information is obtained about the chromophore distribution within the samples by analysis of the frequency dependence of the spectra giving a 3-D image of the posterior ocular tissues.

Complete amino acid sequence of γ-subunit of the GTP-binding protein from cattle retina

The complete amino acid sequence of the γ-subunit of the GTP-binding protein from cattle retina has been established. The polypeptide chain of the γ-subunit consists of 69 amino acid residues and contains the unusual sequence Cys35-Cys36. The M rof the γ-subunit is 8008.7.

Properties of an interphotoreceptor retinoid-binding protein from bovine retina.

Washes and extracts of frozen and fresh cattle retina contain a water-soluble high-molecular-weight, retinoid-binding protein that is distinct from three other retinoid-binding proteins previously isolated from this tissue. The protein can be purified to apparent homogeneity from retinal homogenates by a combination of gel filtration, lectin, and ion-exchange chromatography. Overestimation of the protein molecular weight was observed in several systems involving migration of the protein through a porous network. The approximate molecular weight obtained by sodium dodecyl sulfate-polyacrylamide gel electrophoresis was 140,000, a value consistent with those reported by other laboratories. However, a more detailed analysis using the method of Ferguson revealed the protein to behave anomalously relative to several proteins used as sodium dodecyl sulfate-polyacrylamide gel electrophoresis standards. The apparent radius of the native protein, estimated from calibrated gel filtration, corresponded to a globular protein with a molecular weight of 240,000-280,000, suggesting that the protein was a dimer. However, when the molecular weight of native interphotoreceptor retinoid-binding protein (IRBP) was determined by a method with no shape dependence, sedimentation equilibrium, a value of 131,700 +/- 3,900 g/mol, was obtained. Sedimentation equilibrium in a dissociating solvent (6 M guanidine HCl) yielded a molecular weight of the smallest component of 120,100 +/- 2,300 g/mol. The similarity of values for the denatured and native molecular weight by sedimentation equilibrium demonstrates that the protein is a monomer. In further support of this, no evidence for a dimer was observed in cross-linking experiments with dimethyl suberimidate. The sedimentation coefficient (S0(20),w = 5.73 +/- 0.15 S) and molecular weight from sedimentation equilibrium were employed to calculate the frictional coefficient and Stokes radius of IRBP (f/f0 = 1.64, Rs = 55 A). The high value of f/f0 of the protein provides a reasonable explanation for the over-estimation of the molecular weight of native IRBP on gel filtration. Approximately 2 mol of exogenous all-trans- or 11-cis-retinol were bound per mol of protein (131,000). Approximately 7% of the binding sites were saturated with endogenous ligand (11-cis-retinol, 88%; all-trans-retinol, 12%) following isolation from partially bleached cattle eyes.(ABSTRACT TRUNCATED AT 400 WORDS)

Proteinase-treated photoreceptor discs. Photoelectric activity of the partially-digested rhodopsin and membrane orientation.

Photoreceptor discs from rod outer segments of cattle retina were treated with (a) papain, (b) thermolysin or (c) trypsin, the procedures resulting in the cleavage of the rhodopsin polypeptide chain between (a) 323 and 324, 236 and 237, 241 and 242, (b) 327 and 328, 240 and 241, or (c) 339 and 340 amino acid residues, respectively. In all the cases, partially digested rhodopsins proved to be competent in generating photoelectric potential and increasing membrane conductance of the discs adsorbed onto phospholipid-impregnated collodion film. The kinetics of generation and dissipation of photopotential as well as of formation of metarhodopsin II and of the light-induced rhodopsin protonation were found to be the same in the partially digested preparations and in the intact one. Incubation of papain-treated or thermolysin-treated discs at pH 6.0 induced formation of inside-out vesicles which, when incorporated into the collodion film, generated an oppositely directed photopotential. Treatment of such vesicles with papain gave rise to further cleavages of the polypeptide localized between 30 and 31, 186 and 187 amino acid residues. One more proteinase-sensitive site, localized between 104 and 105 residues, has been discovered in the inside-out vesicles treated with thermolysin. This fact consistent with the scheme of the 'seven column' arrangement of the visual rhodopsin [Ovchinnikov, Yu. A. (1982) FEBS Lett. 148, 179-191]. Rhodopsin, when treated with papain on both sides, was deprived of sixty amino acid residues being split in two sites in the middle part of the polypeptide, but was still active as a photoelectric energy transducer. The main specific feature inherent in the photoelectric response of the papain-treated or thermolysin-treated rhodopsin and absent from the native protein is that the former survives addition of long trains of saturating flashes when the response of the intact preparation becomes negligible. This effect was shown to be due to conversion of partially digested rhodopsin to a photolytic product that at room temperature lived for minutes even in the presence of NH2OH. A 532-nm laser flash effectively converted this product back to rhodopsin.

Properties of an interphotoreceptor retinoid-binding protein from cattle retina

Properties of an interphotoreceptor retinoid-binding protein from cattle retina

Identification of the endogenous retinoids associated with three cellular retinoid-binding proteins from bovine retina and retinal pigment epithelium.

The endogenous retinoids associated with three cellular retinoid-binding proteins from bovine retina and retinal pigment epithelium have been identified by their spectral characteristics and their co-migration with authentic retinoids on high performance liquid chromatography. All-trans-retinol is the only retinoid associated with the cellular retinol-binding protein from retina and retinal pigment epithelium. The saturation of the binding site with native ligand is 0.95 +/- 0.05 mol of retinoid/mol of protein for cellular retinol-binding protein purified from frozen and fresh retina and retinal pigment epithelium. All-trans-retinoic acid has been identified as the endogenous ligand associated with the cellular retinoic acid-binding protein of cattle retina. The saturation of the binding site of cellular retinoic acid-binding protein from frozen and fresh retina (the protein is absent in extracts of retinal pigment epithelium) is about 0.2 mol of retinoid/mol of protein, probably a minimum value due to losses of the ligand. Retinoic acid has not been detected previously in retina. Cellular retinaldehyde-binding protein from retina purifies with two endogenous ligands which cochromatograph on high performance liquid chromatography with 11-cis-retinaldehyde and 11-cis-retinol and occur in a ratio of approximately 3:1, respectively. The binding site of cellular retinaldehyde-binding protein from frozen retina is nearly fully occupied with these two ligands (saturation greater than 0.9 mol of retinoids/mol of protein). In contrast, purified cellular retinaldehyde-binding protein from retinal pigment epithelium carries only 11-cis-retinaldehyde as an endogenous ligand. The saturation of the binding site with this ligand is greater than 0.95.

On the Origin and the Signal-Shaping Mechanism of the Fast Photosignal in the Vertebrate Retina

Fast photosignals (FPS) with R(1) and R(2) components were measured in retinas of cattle, rat, and frog within a temperature range of 0 degrees to 60 degrees C. Except for temperatures near 0 degrees C the signal rise of the R(1) component was determined by the duration of the exciting flash. The kinetics of the R(2) component and the meta transition of rhodopsin in the cattle and rat retina were compared. For the analysis of the FPS it is presupposed that the signal is produced by light-induced charges on the outer segment envelope membrane that spread onto the whole plasma membrane of the photoreceptor cell. To a good approximation, this mechanism can be described by a model circuit with two distinct capacitors. In this model, the charging capacitance of the pigmented outer segment envelope membrane and the capacitance of the receptor's nonpigmented plasma membrane are connected via the extra- and intracellular electrolyte resistances. The active charging is explained by two independent processes, both with exponential rise (R(1) and R(2)), that are due to charge displacements within the pigmented envelope membrane. The time constant tau(2) of the R(2) membrane charging process shows a strong temperature dependence that of the charge redistribution, tau(r), a weak one. In frog and cattle retinas the active charging is much slower within a large temperature range than the passive charge redistribution. From the two-capacitor model it follows for tau(r) << tau(2) that the rise of the R(2) component is determined by tau(r), whereas the decay is given by tau(2). For the rat retina, however, tau(2) approaches tau(r) at physiological temperatures and becomes <tau(r) above 45 degrees C. In this temperature range where tau(2) approximately tau(r), both processes affect rise and decay of the photosignal. The absolute values of tau(r) are in good accordance with the known electric parameters of the photoreceptors. At least in the cattle retina, the time constant tau(2) is identical with that of the slow component of the meta II formation. The strong temperature dependence of the meta transition time gives rise to the marked decrease of the R(2) amplitude with falling temperature. As the R(1) rise could not be fully time resolved the signal analysis does not yield the time constant tau(1) of the R(1) generating process. It could be established, however, within the whole temperature range that the decay of the R(1) component is determined by tau(r). Using an extended model that allows for membrane leakage, we show that in normal ringer solution the membrane time constant does not influence the signal time-course and amplitude.

Light-induced transmission changes in isolated vertebrate retinas

Changes in the transmission of infrared light by visible light stimulus were measured in isolated chicken and cattle retinas. Chicken retinas showed a 1st increasing phase, a reducing phase, and a 2nd increasing phase. Cattle retinas lacked the 1st increasing phase. For chicken retinas, hyperosmolarity suppressed the reducing phase and a lack of glucose suppressed the 2nd increasing phase. Presumably, outer segment discs shrink due to light stimulus, thereafter swelling by utilizing energy from inner segments.

On the rise time of the R1-component of the "early receptor potential": evidence for a fast light-induced charge separation in rhodopsin.

The rising phase of the R1-component of the early receptor potential from isolated cattle retinas was measured with high time resolution. When the measuring capacitance was 133 pF, a latency of about 200 ns was observed. A rise time of about 0.8 mus at 0 degrees C and 1.6 mus at 37 degrees C (extrapolated to ideal measuring conditions) was found. The negative temperature dependence indicates that the rise is not directly related to the production and decay of photolysis products of rhodopsin since the latter have positive temperature coefficients. An increase of the external measuring capacitance caused a slower rise time. The analysis of this effect allowed the determination of the source impedance of the R1-component. The experimental results can be described with a model in which it is assumed that a fast charge separation (ns or ps) takes place in the outer segment of a photoreceptor cell, and spreads passively to the inner segment via the resistance of the interconnecting cilium. The "inner" relaxation could be circumvented by using isolated rod outer segments which lack the passive inner segments, i.e., a rise time of 90 ns could be measured when isolated rod outer segments were attached to Millipore filters. The results suggest that the molecular event leading to the R1-component is an early charge separation which may be as fast as the cis-trans isomerization of the retinal chromophore.

Isolation procedure for bovine retinal rod outer segments.

An isolation procedure to obtain rod outer segments from cattle retinas is reported. Centrifugation of homogenates in discontinuous and continuous sucrose density gradients yields purified photoreceptor cell outer segments. Assay of the final preparation for rhodopsin content gives a ratio of 2.4 for DO280nm/DO498nm.