Synaptic Degeneration of Retinal Ganglion Cells in a Rat Ocular Hypertension Glaucoma Model

Aims Glaucoma is a common neurodegenerative disease that affects retinal ganglion cells (RGCs) and their axons. Little is known of the synaptic degeneration involved in the pathophysiology of glaucoma. Here we used an experimental ocular hypertension model in rats to investigate this issue. Methods Elevated intraocular pressure (IOP) was induced by laser coagulation of the episcleral and limbal veins. RGCs were retrogradely labeled with Fluoro-Gold (FG). The c-fos protein was used as a neuronal connectivity marker. Expression of c-fos in the retinas was investigated by immunohistochemistry at 5 days and 2 weeks after the induction of ocular hypertension. Both surviving RGCs as revealed by retrograde FG-labeled and c-fos-labeled RGCs were counted. Results The c-fos protein was mainly expressed in the nuclei and nucleoli of cells in the ganglion cell layer and inner nuclear layer in the normal retina. We also confirmed that c-fos was also expressed in the nuclei and nucleoli of RGCs retrogradely labeled with FG. There was no significant RGC loss at 5 days but about 13% RGC loss at 2 weeks after the induction of ocular hypertension. The number of RGCs expressing c-fos was significantly lower in the experimental animals at both 5 days and 2 weeks than normal. Conclusion Our study suggests that there is synaptic disconnection for RGCs after ocular hypertension and it may precede the cell death in the early stage. It may provide insight into novel therapeutic strategies to slow the progress of glaucoma. Qing-ling Fu and Xin Li contributed equally to this work.     

Colour vision and visual ecology of the blue-spotted maskray, <Emphasis Type="Italic">Dasyatis kuhlii</Emphasis> Müller &; Henle, 1814

Relatively little is known about the physical structure and ecological adaptations of elasmobranch sensory systems. In particular, elasmobranch vision has been poorly studied compared to the other senses. Virtually nothing is known about whether elasmobranchs possess multiple cone types, and therefore the potential for colour vision, or how the spectral tuning of their visual pigments is adapted to their different lifestyles. In this study, we measured the spectral absorption of the rod and cone visual pigments of the blue-spotted maskray, Dasyatis kuhlii, using microspectrophotometry. D. kuhlii possesses a rod visual pigment with a wavelength of maximum absorbance (λ_max) at 497 nm and three spectrally distinct cone types with λ_max values at 476, 498 and 552 nm. Measurements of the spectral transmittance of the ocular media reveal that wavelengths below 380 nm do not reach the retina, indicating that D. kuhlii is relatively insensitive to ultraviolet radiation. Topographic analysis of retinal ganglion cell distribution reveals an area of increased neuronal density in the dorsal retina. Based on peak cell densities and using measurements of lens focal length made using laser ray tracing and sections of frozen eyes, the estimated spatial resolving power of D. kuhlii is 4.10 cycles per degree.     

A Laser-Induced Mouse Model with Long-Term Intraocular Pressure Elevation

Purpose

To develop and characterize a mouse model with intraocular pressure (IOP) elevation after laser photocoagulation on the trabecular meshwork (TM), which may serve as a model to investigate the potential of stem cell-based therapies for glaucoma.

Methods

IOP was measured in 281 adult C57BL/6 mice to determine normal IOP range. IOP elevation was induced unilaterally in 50 adult mice, by targeting the TM through the limbus with a 532-nm diode laser. IOP was measured up to 24 weeks post-treatment. The optic nerve damage was detected by electroretinography and assessed by semiautomatic counting of optic nerve axons. Effects of laser treatment on the TM were evaluated by histology, immunofluorescence staining, optical coherence tomography (OCT) and transmission electron microscopy (TEM).

Results

The average IOP of C57BL/6 mice was 14.5±2.6 mmHg (Mean ±SD). After laser treatment, IOP averaged above 20 mmHg throughout the follow-up period of 24 weeks. At 24 weeks, 57% of treated eyes had elevated IOP with the mean IOP of 22.5±2.5 mmHg (Mean ±SED). The difference of average axon count (59.0%) between laser treated and untreated eyes was statistically significant. Photopic negative response (PhNR) by electroretinography was significantly decreased. CD45+ inflammatory cells invaded the TM within 1 week. The expression of SPARC was increased in the TM from 1 to 12 weeks. Histology showed the anterior chamber angle open after laser treatment. OCT indicated that most of the eyes with laser treatment had no synechia in the anterior chamber angles. TEM demonstrated disorganized and compacted extracellular matrix in the TM.

Conclusions

An experimental murine ocular hypertension model with an open angle and optic nerve axon loss was produced with laser photocoagulation, which could be used to investigate stem cell-based therapies for restoration of the outflow pathway integrity for ocular hypertension or glaucoma.     

康柏西普与雷珠单抗对年龄相关性黄斑变性患者血清CRP、VEGF、眼压及视力的影响

目的:探讨康柏西普与雷珠单抗对年龄相关性黄斑变性(AMD)患者血清中C反应蛋白(CRP)、血管内皮生长因子(VEGF)水平、眼压(IOP)及视力的影响。方法:选取2015年4月到2016年5月在我院接受治疗的AMD患者70例作为研究对象,采用随机数字表法将所有患者分为观察组和对照组各35例,所有患者给予内眼手术标准玻璃体注射治疗,观察组给予康柏西普注射液1.5mg腔内注射,对照组给予雷珠单抗注射液0.5 mL腔内注射,对比两组患者治疗前、治疗后3d血清CRP、VEGF水平;记录并对比两组患者视力、IOP水平及并发症情况。结果:两组患者治疗后3d血清CRP、VEGF水平较治疗前下降,且观察组患者CRP、VEGF水平低于对照组(P0.05);两组患者治疗后3d视力最小视角对数(logMAR)及IOP较治疗前降低,且观察组患者视力logMAR及IOP水平明显低于对照组,差异均有统计学意义(P0.05)。治疗后3d,两组各出现1例玻璃体出血,并发症发生率比较差异无统计学意义(P0.05)。结论:相对于雷珠单抗,康柏西普更能有效降低AMD患者血清CRP、VEGF水平和IOP,提高患者视力,改善病情,值得临床推广应用。     

Methylnitrosourea (MNU)-induced Retinal Degeneration and Regeneration in the Zebrafish: Histological and Functional Characteristics

Retinal degenerative diseases, e.g. retinitis pigmentosa, with resulting photoreceptor damage account for the majority of vision loss in the industrial world. Animal models are of pivotal importance to study such diseases. In this regard the photoreceptor-specific toxin N-methyl-N-nitrosourea (MNU) has been widely used in rodents to pharmacologically induce retinal degeneration. Previously, we have established a MNU-induced retinal degeneration model in the zebrafish, another popular model system in visual research.A fascinating difference to mammals is the persistent neurogenesis in the adult zebrafish retina and its regeneration after damage. To quantify this observation we have employed visual acuity measurements in the adult zebrafish. Thereby, the optokinetic reflex was used to follow functional changes in non-anesthetized fish. This was supplemented with histology as well as immunohistochemical staining for apoptosis (TUNEL) and proliferation (PCNA) to correlate the developing morphological changes.In summary, apoptosis of photoreceptors occurs three days after MNU treatment, which is followed by a marked reduction of cells in the outer nuclear layer (ONL). Thereafter, proliferation of cells in the inner nuclear layer (INL) and ONL is observed. Herein, we reveal that not only a complete histological but also a functional regeneration occurs over a time course of 30 days. Now we illustrate the methods to quantify and follow up zebrafish retinal de- and regeneration using MNU in a video-format.     

Measuring Spinal Presynaptic Inhibition in Mice By Dorsal Root Potential Recording In Vivo

Presynaptic inhibition is one of the most powerful inhibitory mechanisms in the spinal cord. The underlying physiological mechanism is a depolarization of primary afferent fibers mediated by GABAergic axo-axonal synapses (primary afferent depolarization). The strength of primary afferent depolarization can be measured by recording of volume-conducted potentials at the dorsal root (dorsal root potentials, DRP). Pathological changes of presynaptic inhibition are crucial in the abnormal central processing of certain pain conditions and in some disorders of motor hyperexcitability. Here, we describe a method of recording DRP in vivo in mice. The preparation of spinal cord dorsal roots in the anesthetized animal and the recording procedure using suction electrodes are explained. This method allows measuring GABAergic DRP and thereby estimating spinal presynaptic inhibition in the living mouse. In combination with transgenic mouse models, DRP recording may serve as a powerful tool to investigate disease-associated spinal pathophysiology. In vivo recording has several advantages compared to ex vivo isolated spinal cord preparations, e.g. the possibility of simultaneous recording or manipulation of supraspinal networks and induction of DRP by stimulation of peripheral nerves.     

兔先天性青光眼网络膜血管改变

目的　研究青光眼对视网膜脉络膜血液循环的影响。方法　选24月龄、体重3.5～4kg的先天性青光眼大耳白兔5只(7只眼),选10只同龄大耳白兔作为对照组。另选10只2月龄、体重2kg大耳白兔前房内灌注生理盐水制成急性高眼压模型。对三组兔进行眼底照像、闪光视诱发电位(FVEP)检查,观察视网膜脉络膜血管形态和FVEP的变化。对人工急性高眼压组还进行了闪光视网膜电流图(FERG)检查。结果　先天性青光眼组与同龄对照组相比视网膜脉络膜末梢血管网明显减少;人工急性高眼压组眼压升高后首先使视网膜脉络膜末梢血管网灌流不足,随着眼压的继续升高脉络膜大血管变细,末梢血管网灌流不足加重,眼压极度升高时脉络膜大血管血流中断。同龄正常对照组的FVEP的主波P100潜伏期是(83±9)ms,先天性青光眼组则为(112±14)ms,差异有非常显著意义(P＜0.01);人工急性高眼压组高眼压前为(69±5)ms,眼压60～80mm　Hg时延长为(81±7)ms,眼压在100～130mmHg时FVEP波形低平,近似直线;眼压恢复正常后2hFVEP的P100潜伏期为(82±8)ms。人工急性高眼压前后FERG变化显著。结论　青光眼可以影响视网膜脉络膜血液循环;可使FVEP、FERG发生变化。     

汉防己甲素滴眼液对高眼压模型大鼠的降眼压作用

目的比较观察汉防己甲素滴眼液与0.5%噻吗心安滴眼液对高眼压模型大鼠及正常大鼠降眼压的作用。方法正常SD大鼠共分4组：不同浓度的汉防己甲素滴眼液组（0.1%、0.2%、0.3%）及阳性对照组0.5%噻吗心安,药物滴右眼各一滴,阴性对照组生理盐水滴左眼、测量滴药前24h和滴药后1、3、6、24、48、72h的眼压。应用倍频532激光对SD大鼠右眼上巩膜静脉以及小梁网所在区域实施光凝术建立高眼压大鼠模型。高眼压模型鼠共分5组：不同浓度的汉防己甲素滴眼液0.05%、0.1%、0.2%、0.3%及阳性对照组0.5%噻吗心安,右眼即模型眼滴用药物,左眼作为空白对照。测量术前后的眼压。结果汉防己甲素滴眼液对大鼠正常眼压无降压作用（P〉0.05）。对高眼压大鼠用药后24h、72h、1周后,0.3%汉防己甲素滴眼液组降低眼压的幅度与0.5%噻吗心安滴眼液降低眼压的幅度相似（P〉0.05）;0.05%、0.1%、0.2%汉防己甲素滴眼液组也有明显的降压作用,但与0.5%噻吗心安滴眼液相比,降压幅度低于后者（P〈0.05）。结论0.05%、0.1%、0.2%、0.3%汉防己甲素滴眼液均有降低大鼠高眼压的作用,其中0.3%浓度的汉防己甲素滴眼液降眼压效果与0.5%的噻吗心安类似。汉防已甲素滴眼液作为一种治疗青光眼的药物有着良好应用前景。     

内皮抑素对眼部新生血管抑制的研究进展

眼部新生血管存在于多种常见的眼病的发展过程中,对视功能危害大,是致盲的主要原因之一。包括糖尿病视网膜病变,视网膜栓塞,早产儿视网膜病变,老年性黄斑变性等眼病。由于其发病机制尚未完全清楚,因此目前仍无确切有效的药物治疗方法。内皮抑素（Endostatin,ES）是1997年首先从小鼠血管内皮瘤EOMA细胞培养上清中发现的,是胶原xⅧ的蛋白降解产物,分子质量约为20KD,为胶原xⅧc端非胶原区（NCl）内的184个氨基酸片段。ES是目前发现的最强的血管生成抑制因子,可抑制VEGF,bFGF,EGF等刺激的血管内皮细胞的增殖和迁移,诱导其凋亡,进而抑制新生血管的形成。通过抑制眼部新生血管的实验研究表明,ES是当前抗新生血管疗法中最有潜力的一种新药。本文就内皮抑素的结构特点及其对眼部新生血管的治疗研究进展作一综述。     

内皮抑素对眼部新生血管抑制的研究进展

眼部新生血管存在于多种常见的眼病的发展过程中,对视功能危害大,是致盲的主要原因之一。包括糖尿病视网膜病变,视网膜栓塞,早产儿视网膜病变,老年性黄斑变性等眼病。由于其发病机制尚未完全清楚,因此目前仍无确切有效的药物治疗方法。内皮抑素(Endostatin,ES)是1997年首先从小鼠血管内皮瘤EOMA细胞培养上清中发现的,是胶原XⅧ的蛋白降解产物,分子质量约为20KD,为胶原XⅧC端非胶原区(NC1)内的184个氨基酸片段。ES是目前发现的最强的血管生成抑制因子,可抑制VEGF,bFGF,EGF等刺激的血管内皮细胞的增殖和迁移,诱导其凋亡,进而抑制新生血管的形成。通过抑制眼部新生血管的实验研究表明,ES是当前抗新生血管疗法中最有潜力的一种新药。本文就内皮抑素的结构特点及其对眼部新生血管的治疗研究进展作一综述。     

Rescue of Retinal Function by BDNF in a Mouse Model of Glaucoma

Vision loss in glaucoma is caused by progressive dysfunction of retinal ganglion cells (RGCs) and optic nerve atrophy. Here, we investigated the effectiveness of BDNF treatment to preserve vision in a glaucoma experimental model. As an established experimental model, we used the DBA/2J mouse, which develops chronic intraocular pressure (IOP) elevation that mimics primary open-angle glaucoma (POAG). IOP was measured at different ages in DBA/2J mice. Visual function was monitored using the steady-state Pattern Electroretinogram (P-ERG) and visual cortical evoked potentials (VEP). RGC alterations were assessed using Brn3 immunolabeling, and confocal microscope analysis. Human recombinant BDNF was dissolved in physiological solution (0.9% NaCl); the effects of repeated intravitreal injections and topical eye BDNF applications were independently evaluated in DBA/2J mice with ocular hypertension. BDNF level was measured in retinal homogenate by ELISA and western blot. We found a progressive decline of P-ERG and VEP responses in DBA/2J mice between 4 and 7 months of age, in relationship with the development of ocular hypertension and the reduction of Brn3 immunopositive RGCs. Conversely, repeated intravitreal injections (BDNF concentration = 2 µg/µl, volume = 1 µl, for each injection; 1 injection every four days, three injections over two weeks) and topical eye application of BDNF eye-drops (12 µg/µl, 5 µl eye-drop every 48 h for two weeks) were able to rescue visual responses in 7 month DBA/2J mice. In particular, BDNF topical eye treatment recovered P-ERG and VEP impairment increasing the number of Brn3 immunopositive RGCs. We showed that BDNF effects were independent of IOP reduction. Thus, topical eye treatment with BDNF represents a promisingly safe and feasible strategy to preserve visual function and diminish RGC vulnerability to ocular hypertension.     

Insulin-like growth factor-1 binding protein 3 (IGFBP-3) promotes recovery from trauma-induced expression of inflammatory and apoptotic factors in retina

Ocular trauma affects 20% of Americans in their lifetime and can cause permanent visual system damage. We have used a mouse model of ocular trauma (exposure to an air blast from a paintball gun) to examine pathways that trigger the resulting retinal damage and to develop treatment strategies that might ameliorate the deleterious effects of trauma on retinal tissue. Our previous studies have shown that ocular blast causes an increase in protein levels of inflammatory mediators and apoptotic factors, including tumor necrosis factor alpha (TNFα) and interleukin-1-beta (IL-1β), as well as the apoptotic markers, Bax, cytochrome C, and cleaved caspase 3. Furthermore, topical treatment by eye drop application of a β-adrenergic receptor agonist, Compound 49b, was shown to decrease these inflammation/apoptosis markers and thus ameliorate the effects of blast trauma. We postulate that the protective effect of Compound 49b may be linked to its demonstrated ability to activate the β-adrenergic receptor and in turn trigger production of insulin-like growth factor binding protein 3 (IGFBP-3). In the current study, we tested this hypothesis using mice with minimal IGFBP-3 activity (IGFBP-3 knockdown mouse) vs. wildtype mice. We found that ocular blast alone did not affect IGFBP-3 levels in retinas of wild type or knockdown mice and surprisingly, the lower levels of IGFBP-3 in knockdown animals did not exacerbate the blast-induced increase in protein levels of inflammation/apoptosis markers. Nevertheless, the levels of IGFBP-3 were significantly increased in knockdown mouse retina by treatment with Compound 49b 24 h post-trauma and as expected, the increase in IGFBP-3 was linked to a decrease in inflammation/apoptosis markers. We conclude that while lowered IGFBP-3 may not make the retina more vulnerable to blast injury, an increase in IGFBP-3 post-trauma may play an important role in limiting trauma-induced inflammatory and apoptotic pathways leading to retinal damage. Eye drop application of the β-adrenergic receptor agonist, Compound 49b, provides a promising treatment strategy for increasing IGFBP-3 levels to promote recovery from retinal inflammation and apoptosis after ocular blast.     

Effects of intravitreous sodium hydrosulfide on intraocular pressure and retinopathy in ocular hypertensive rats

We investigated the possible neuroprotectant and intraocular pressure (IOP) lowering effects of intravitreous injection of sodium hydrosulfide (NaSH) in a rodent model of experimental glaucoma. Glaucoma currently is treated by controlling IOP using medications and/or surgery. These methods are not entirely adequate for all patients. We divided 24 rats into three groups. For the control group, the right eye was treated with intravitreous saline. For the glaucoma group, ocular hypertension was induced by photocoagulating three episcleral veins and the limbal plexus of the right eye using an argon laser, then saline was injected into the vitreous of these eyes during the third week. For the NaSH group, rats were treated with intravenous NaSH 3 weeks after photocoagulation. IOP was measured each week during the 6 week experimental period. Coagulating the episcleral veins rapidly increased the IOP of rat eyes. Intravitreous injection of NaSH significantly reduced IOP. Intravitreous NaSH prevented degeneration of the retina and decreased the number of apoptotic cells. Intravitreous NaSH appeared to reduce IOP and to prevent IOP induced retinopathy in rats.     

Murine neonatal infection provides an efficient model for congenital ocular toxoplasmosis

Congenital infection is one of the most serious settings of infection with the apicomplexan parasite Toxoplasma gondii. Ocular diseases, such as retinochoroiditis, are the most common sequels of such infection in utero. However, while numerous studies have investigated the physiopathology of acquired toxoplasmosis, congenital infection has been largely neglected so far. Here, we establish a mouse model of congenital ocular toxoplasmosis. Parasite load and ocular pathology have been followed for the first 4 weeks of life. Ocular infection developed slowly compared to cerebral infection. Even after 4 weeks, not all eyes were infected and ocular parasite load was low. Therefore, we evaluated a scheme of neonatal infection to overcome problems associated with congenital infection. Development of infection and physiopathology was similar, but at a higher, more reliable rate. In summary, we have established a valuable model of neonatal ocular toxoplasmosis, which facilitates the research of the underlying physiopathological mechanisms and new diagnostic approaches of this pathology.     

The Short-Term Effects of Exercise on Intraocular Pressure,Choroidal Thickness and Axial Length

Purpose

To explore ocular changes in healthy people after exercise.

Methods

Twenty five volunteers underwent exercise for 15 minutes on a treadmill. Measurements of choroidal thickness, intraocular pressure (IOP), ocular biometry, and blood pressure were taken before and after exercise. Enhanced Depth Imaging optical coherence tomography (EDI-OCT) was used to measure choroidal thickness at the fovea. Intraocular pressure (IOP) was measured by Goldmann applanation tonometry. Ocular biometric measures were collected using A scan ultrasound. Blood pressure was measured concurrently with the acquisition of the scans.

Results

Twenty five volunteers (25 eyes) with a mean age of 25.44±3.25 years were measured. There was a significant increase in systolic and diastolic pressure after exercise (P<0.05). The IOP showed a significant decrease after exercise (P<0.05). However there was no significant difference in the mean choroidal thickness, ocular axial length, anterior chamber depth, lens thickness, or vitreous length before and after exercise measurements (P>0.05).

Conclusion

There was a significant decrease in IOP from exercise without a change in choroidal thickness and ocular biometric measures. IOP and choroidal thickness were not correlated, suggesting that the IOP decrease from exercise is not due to changes in choridal thickness.     

Localization of visual pigments within rhabdoms of the compound eye of Spodoptera exempta (Insecta,Noctuidae)

Summary In the noctuid moth Spodoptera exempta, the distribution of visual pigments within the fused rhabdoms of the compound eyes was investigated by electron microscopy. Each ommatidium regularly contains eight receptor cells belonging to three morphological types: one distal, six medial, and one basal cell (Meinecke 1981); four different visual pigments — absorption maxima at approximately 355, 465, 515, and 560 nm — are known to occur within the eye (Langer et al. 1979). The compound eyes were illuminated in situ by use of monochromatic light of different wavelengths. This illumination produced a wide scale of structural changes in the microvilli of the rhabdomeres of individual cells. Preparation of eyes by freeze-substitution revealed the structural changes in the rhabdomeres to be effects of light occurring in vivo.The degree of structural changes may be considerably different in rhabdomeres within the same ommatidium; it was found to depend on the wavelength and the duration of illumination, the intensity received by the ommatidia as well as the spectral sensitivity of the receptor cells. Therefore, it was possible to estimate the spectral sensitivities of the morphological types of receptor cells. Generally, all medial cells are green receptors and all basal cells red receptors; distal cells are blue receptors in about two-thirds of the ommatidia, while in the remaining third of them distal cells are sensitive to ultraviolet light.Supported by Deutsche Forschungsgemeinschaft, Sonderforschungsbereich 114 (Bionach)     

Changes in Glial Fibrillary Acidic Protein (GFAP) Immonureactivity Reflect Neuronal States

The astroglial marker, glial fibrillary acidic protein (GFAP) was investigated by immunohistochemistry in various brain areas in order to see its fluctuations in various functional states. Different neuronal states were either experimentally induced or studied under physiological conditions. To produce experimental alterations the visual system was chosen as a model. Upon lesioning of the lateral geniculate body with the stereotaxic injection of ibotenic acid an increase in GFAP immunoreactivity could be induced in layers III and IV of the ipsilateral visual cortex where geniculo-cortical fibres terminate. Electron microscopy has revealed a synchronous degeneration of synaptic terminals and the hypertrophy of perisynaptic astrocyte processes. To study changes in the intact animal the effect of illumination was observed. In the lateral geniculate body the dorsal subnucleus was found immunonegative when studied at day and positive at night. Similar changes were observed in the suprachiasmatic nucleus. As to more generalized influences, the effect of gonadal steroids on the GFAP-reaction interpeduncular nucleus, an area not involved in hormonal regulatory mechanisms was studied. In males only castration could reduce constantly high GFAP immonoreactivity, whereas in females GFAP showed wide-range sexual cycle-related fluctuations. It was concluded that changes in GFAP immunoreactivity can indicate synaptic events whithin a circumscribed area of the brain. Special issue article in honor of Dr. Ricardo Tapia.     

Assessment of thermal dehydration using the human eye: What is the potential?

Human hydration assessment is a key component for the prevention and proper treatment of heat-related fluid and electrolyte imbalances within military, sports and clinical medicine communities. Despite the availability of many different methods for assessing hydration status, the need for a valid method or technology that is simple, rapid, non-invasive, universal (detects both hypertonic and isotonic hypovolaemia) and is applicable for static (single point in time) and dynamic (change across time) hydration assessment is widely acknowledged. The eye is one candidate body region that might afford such a measure given the intricate balance between ocular dynamics (tear and aqueous humor formation) and blood (plasma osmolality and volume), which is considered the criterion measure for hydration assessment. The aim of this review is to introduce and discuss the potential for using ocular measurements for non-invasive hydration assessment, including tear fluid osmolarity (Tosm), non-invasive tear break-up time (NITBUT) and intraocular pressure (IOP). There is a relevant physiological basis for testing the merit of ocular measures for human hydration assessment and recent data indicate that Tosm and IOP may have utility. Further investigations are warranted to determine the degree to which ocular measures can act as accurate and reliable non-invasive hydration status markers.     

Genetic modifiers of cardiovascular phenotype caused by elastin haploinsufficiency act by extrinsic noncomplementation

Elastin haploinsufficiency causes the cardiovascular complications associated with Williams-Beuren syndrome and isolated supravalvular aortic stenosis. Significant variability exists in the vascular pathology in these individuals. Using the Eln(+/-) mouse, we sought to identify the source of this variability. Following outcrossing of C57Bl/6J Eln(+/-), two backgrounds were identified whose cardiovascular parameters deviated significantly from the parental strain. F1 progeny of the C57Bl/6J; Eln(+/-)x129X1/SvJ were more hypertensive and their arteries less compliant. In contrast, Eln(+/-) animals crossed to DBA/2J were protected from the pathologic changes associated with elastin insufficiency. Among the crosses, aortic elastin and collagen content did not correlate with quantitative vasculopathy traits. Quantitative trait locus analysis performed on F2 C57; Eln(+/-)x129 intercrosses identified highly significant peaks on chromosome 1 (LOD 9.7) for systolic blood pressure and on chromosome 9 (LOD 8.7) for aortic diameter. Additional peaks were identified that affect only Eln(+/-), including a region upstream of Eln on chromosome 5 (LOD 4.5). Bioinformatic analysis of the quantitative trait locus peaks revealed several interesting candidates, including Ren1, Ncf1, and Nos1; genes whose functions are unrelated to elastic fiber assembly, but whose effects may synergize with elastin insufficiency to predispose to hypertension and stiffer blood vessels. Real time RT-PCR studies show background-specific increased expression of Ncf1 (a subunit of the NOX2 NAPDH oxidase) that parallel the presence of increased oxidative stress in Eln(+/-) aortas. This finding raises the possibility that polymorphisms in genes affecting the generation of reactive oxygen species alter cardiovascular function in individuals with elastin haploinsufficiency through extrinsic noncomplementation.