Karolina Motloch

Brain and Retinal Pericytes: Origin, Function and Role

Pericytes are specialized mural cells located at the abluminal surface of capillary blood vessels, embedded within the basement membrane. In the vascular network these multifunctional cells fulfil diverse functions, which are indispensable for proper homoeostasis. They serve as microvascular stabilizers, are potential regulators of microvascular blood flow and have a central role in angiogenesis, as they for example regulate endothelial cell proliferation. Furthermore, pericytes, as part of the neurovascular unit, are a major component of the blood-retina/brain barrier. CNS pericytes are a heterogenic cell population derived from mesodermal and neuro-ectodermal germ layers acting as modulators of stromal and niche environmental properties. In addition, they display multipotent differentiation potential making them an intriguing target for regenerative therapies. Pericyte-deficiencies can be cause or consequence of many kinds of diseases. In diabetes, for instance, pericyte-loss is a severe pathological process in diabetic retinopathy (DR) with detrimental consequences for eye sight in millions of patients. In this review, we provide an overview of our current understanding of CNS pericyte origin and function, with a special focus on the retina in the healthy and diseased. Finally, we highlight the role of pericytes in de- and regenerative processes.

Intraocular Pressure Course During the Femtosecond Laser-Assisted Cataract Surgery in Porcine Cadaver Eyes

Purpose Femtosecond laser-assisted cataract surgery (FLACS) is an increasingly common procedure. Most laser systems require corneal applanation and thereby increase intraocular pressure (IOP). The purpose of the present study was to evaluate the IOP changes that occur during the FLACS procedure performed using the Catalys femtosecond laser system. Methods IOP was measured by direct cannulation of the vitreous body of porcine cadaver eyes (N = 20). By inserting a second cannula connected to a water column, all the eyes were set to a baseline IOP of 20 mm Hg. The eyes were lifted by custom-made stands to achieve the appropriate height and position under the Catalys system. The standard FLACS procedure was performed using varying fragmentation times to assess the influence of tissue fragmentation times on IOP peaks. Results We identified significant IOP elevations from baseline IOP levels during all steps of the FLACS procedure (baseline: 20.28 ± 1.32 mm Hg; vacuum: 34.48 ± 4.21 mm Hg; capture: 47.90 ± 13.02 mm Hg; lock: 48.41 ± 9.04 mm Hg; analysis: 47.15 ± 5.97 mm Hg; capsulotomy: 45.74 ± 6.52 mm Hg; fragmentation: 48.41 ± 6.80 mm Hg; end: 17.81 ± 1.61 mm Hg; all P < 0.001). Furthermore, the tissue fragmentation time had a significant effect on the peak IOP values detected (R = 0.62, P = 0.04, n = 9). Conclusions The present study reveals significant IOP increases during FLACS procedures carried out using the Catalys system.

Through modulation of cardiac Ca2+‐handling, UCP2 affects cardiac electrophysiology and influences the susceptibility for Ca2+‐mediated arrhythmias

What is the central question of this study? Knockdown of UCP2 reduces mitochondrial Ca2+ uptake. This suggests that Ucp2 knockout mice need to have additional effects on cytosolic Ca2+ handling to prevent Ca2+ overload. However, the specific mechanisms and their impact on cardiac electrophysiology remain speculative. What is the main finding and its importance? In Ucp2 knockout mice, decreased mitochondrial Ca2+ uptake is compensated for by functional inhibition of L‐type Ca2+ channels and resultant shortening of action potential duration. UCP2‐dependent modulations have a major impact on cardiac electrophysiology, resulting in alterations of ECG characteristics and a higher susceptibility to Ca2+‐mediated ventricular arrhythmias.

Lymphatic and vascular markers in an optic nerve crush model in rat

Abstract Only few tissues lack lymphatic supply, such as the CNS or the inner eye. However, if the scleral border is compromised due to trauma or tumor, lymphatics are detected in the eye. Since the situation in the optic nerve (ON), part of the CNS, is not clear, the aim of this study is to screen for the presence of lymphatic markers in the healthy and lesioned ON. Brown Norway rats received an unilateral optic nerve crush (ONC) with defined force, leaving the dura intact. Lesioned ONs and unlesioned contralateral controls were analyzed 7 days (n = 5) and 14 days (n = 5) after ONC, with the following markers: PDGFRb (pericyte), Iba1 (microglia), CD68 (macrophages), RECA (endothelial cell), GFAP (astrocyte) as well as LYVE‐1 and podoplanin (PDPN; lymphatic markers). Rat skin sections served as positive controls and confocal microscopy in single optical section mode was used for documentation. In healthy ONs, PDGFRb is detected in vessel‐like structures, which are associated to RECA positive structures. Some of these PDGFRb+/RECA+ structures are closely associated with LYVE‐1+ cells. Homogenous PDPN‐immunoreactivity (IR) was detected in healthy ON without vascular appearance, showing no co‐localization with LYVE‐1 or PDGFRb but co‐localization with GFAP. However, in rat skin controls PDPN‐IR was co‐localized with LYVE‐1 and further with RECA in vessel‐like structures. In lesioned ONs, numerous PDGFRb+ cells were detected with network‐like appearance in the lesion core. The majority of these PDGFRb+ cells were not associated with RECA‐IR, but were immunopositive for Iba1 and CD68. Further, single LYVE‐1+ cells were detected here. These LYVE‐1+ cells were Iba1‐positive but PDPN‐negative. PDPN‐IR was also clearly absent within the lesion site, while LYVE‐1+ and PDPN+ structures were both unaltered outside the lesion. In the lesioned area, PDGFRb+/Iba1+/CD68+ network‐like cells without vascular association might represent a subtype of microglia/macrophages, potentially involved in repair and phagocytosis. PDPN was detected in non‐lymphatic structures in the healthy ON, co‐localizing with GFAP but lacking LYVE‐1, therefore most likely representing astrocytes. Both, PDPN and GFAP positive structures are absent in the lesion core. At both time points investigated, no lymphatic structures can be identified in the lesioned ON. However, single markers used to identify lymphatics, detected non‐lymphatic structures, highlighting the importance of using a panel of markers to properly identify lymphatic structures. HighlightsPDPN expression was detected in GFAP positive astrocytes in the rat ON.LYVE‐1 is expressed in cells with microglial/macrophagic activity.Absence of lymphatic structures within the rat ON under physiological conditions.No formation of lymphatic structures within the rat ON following ONC trauma.

Retinal Vessel Diameter Responses to Central Electrical Stimulation in the Rat: Effect of Nitric Oxide Synthase Inhibition.

PURPOSE Recent histological data suggest autonomic innervation of the central retinal artery. In the present study, we investigated the effect of electrical brain stem stimulation at the superior salivatory nucleus (SSN) on the retinal vessel diameter in rats and whether nitric oxide mediates a possible effect. METHODS Sprague-Dawley rats (n = 12) were anesthetized using pentobarbital sodium (50 mg/kg intraperitoneally). The animals were artificially ventilated and the femoral artery and vein were cannulated for blood pressure measurement and drug administration. After a craniotomy was performed, a unipolar stainless steel electrode was inserted into the brainstem at the coordinates of the SSN. Stimulations were performed at 20 Hz, 9 μA, 1 ms pulse duration and 200 pulses. Retinal vessel diameters were measured continuously with the Imedos DVA-R, a noncontact fundus camera for rats with image analysis software. After control measurements, L-NAME, a nonspecific inhibitor of NO synthase, was applied intravenously (10 mg/kg), and the SSN stimulations were repeated. RESULTS Stimulation at the SSN coordinates increased the retinal arterial diameter by 6.41% ± 1.65% and the venous diameter by 3.48% ± 1.93% (both P < 0.05). Application of L-NAME reduced the arterial response significantly to 2.93% ± 0.91%, but did not change the venous response. Mean arterial pressure, carotid blood flow, and heart rate remained unaltered (by the stimulation). CONCLUSIONS The present study demonstrates that the retinal circulation reacts to electric stimulation at the SSN coordinates in rats. Nitric oxide is involved in the response, but it is not the sole neurotransmitter.

Aquaporin expression and localization in the rabbit eye

Aquaporins (AQPs) are important for ocular homeostasis and function. While AQP expression has been investigated in ocular tissues of human, mouse, rat and dog, comprehensive data in rabbits are missing. As rabbits are frequently used model organisms in ophthalmic research, the aim of this study was to analyze mRNA expression and to localize AQPs in the rabbit eye. The results were compared with the data published for other species. In cross sections of New Zealand White rabbit eyes AQP0 to AQP5 were labeled by immunohistology and analyzed by confocal microscopy. Immunohistological findings were compared to mRNA expression levels, which were analyzed by quantitative reverse transcription real time polymerase chain reaction (qRT-PCR). The primers used were homologous against conserved regions of AQPs. In the rabbit eye, AQP0 protein expression was restricted to the lens, while AQP1 was present in the cornea, the chamber angle, the iris, the ciliary body, the retina and, to a lower extent, in optic nerve vessels. AQP3 and AQP5 showed immunopositivity in the cornea. AQP3 was also present in the conjunctiva, which could not be confirmed for AQP5. However, at a low level AQP5 was also traceable in the lens. AQP4 protein was detected in the ciliary non-pigmented epithelium (NPE), the retina, optic nerve astrocytes and extraocular muscle fibers. For most tissues the qRT-PCR data confirmed the immunohistology results and vice versa. Although species differences exist, the AQP protein expression pattern in the rabbit eye shows that, especially in the anterior section, the AQP distribution is very similar to human, mouse, rat and dog. Depending on the ocular regions investigated in rabbit, different protein and mRNA expression results were obtained. This might be caused by complex gene regulatory mechanisms, post-translational protein modifications or technical limitations. However, in conclusion the data suggest that the rabbit is a useful in-vivo model to study AQP function and the effects of direct and indirect intervention strategies to investigate e. g. mechanisms for intraocular pressure modulation or cornea transparency regulation.