Nonvascular Tissues Research Articles

Intraarticular Treatments for Osteoarthritis: New Perspectives

To date, no targeted treatments for osteoarthritis have been developed. Therefore the challenge for the next years is to find a treatment that may slow down the progression of the disease. Osteoarthritis of the weight-bearing joints, such as knee OA, is more a local mechanical driven disease than a generalized one. To reach a non-vascularised tissue such as the cartilage, local intra-articular administration of drugs should be considered. The purpose of this review is to evaluate the advantages of local intra-articular drug administration compared with a systemic one in patients with osteoarthritis of weight-bearing joints. New perspectives of such strategy are reviewed, including anti-cytokine therapy, gene therapy, delivery of growth factors, stem cells therapy and new lubricant agents such as lubricin. The main goal in the future will be to increase the residence time of the drug in the joint while improving its diffusion within the target tissues. One key question will be how to better define the patients likely to benefit by such an approach and when the treatment is most likely to work. Most importantly, the treatment strategy must be selected according to the pattern in an individual patient. Finally, with all intra-articular treatments, the risk/benefit ratio must be carefully evaluated.

Reconstruction of the cranial base after endonasal skull base surgery: Local tissue flaps

Advances in endoscopic techniques and technology have substantially increased the scope of endonasal skull base surgery. Large dural defects after expanded endonasal approaches (EEA) present a challenge for reconstruction and have revealed the limitations of conventional reconstructive methods. Initial reconstructive attempts using nonvascularized free tissue grafts resulted in unacceptably high rates of cerebrospinal fluid fistulae. The advent of vascularized, pedicled flaps has substantially improved these outcomes. These flaps also have the advantage of a large surface area and can be mobilized over a large arc of rotation. Vascularized tissue heals quickly and can tolerate postoperative radiation therapy. We have developed 3 nasal vascularized flaps that can be used for skull base reconstruction after EEA. The anatomy, technique, and reconstructive considerations for the nasoseptal flap, the inferior turbinate flap, and the middle turbinate flap will be reviewed in detail in the following report.

The gene encoding the catalytically inactive β-amylase BAM4 involved in starch breakdown in Arabidopsis leaves is expressed preferentially in vascular tissues in source and sink organs

Genetic studies in Arabidopsis thaliana have shown that two members of the beta-amylase (BAM) family BAM3 and BAM4 are required for leaf starch breakdown at night. Both are plastid proteins and while BAM3 encodes an active BAM, BAM4 is not an active alpha-1,4-glucan hydrolase. To gain further insight into the possible function of BAM4 we constructed reporter genes using promoters for both BAM3 and BAM4 genes, driving beta-glucuronidase (GUS) and luciferase (LUC) expression in transgenic Arabidopsis plants. Both promoters directed expression in vascular tissue throughout the plant including cotyledons, leaves, petioles, stems, petals, siliques and roots. Tissue sections showed expression to be focused in phloem cells in stem and petiole. The BAM3 promoter was also expressed strongly throughout the photosynthetic tissues of leaves, sepals and siliques, whereas the BAM4 promoter was not. Conversely, the BAM4 promoter was active in root tip but the BAM3 promoter was not. To confirm these expression patterns and to compare with expression of other starch genes we carried-out RT-PCR analysis on RNA from vascular (replum) and non-vascular (valve) tissues of siliques. This confirmed that BAM4 expression together with RAM1 (BAM5) and GWD2 genes is stronger in the replum than the valve, whereas BAM3 is strong in both tissues. These results show that even though BAM3 and BAM4 genes apparently interact genetically in leaf starch metabolism, BAM4 is preferentially expressed in non-photosynthetic vascular tissue, so revealing a potentially greater level of complexity in the control of starch breakdown than had previously been recognised.

Regulating the Responses of the Neurovascular Unit to Focal Ischemia

During focal cerebral ischemia, cerebral microvessel and their recipient neurons, as well as other components of the neurovascular unit (NVU) , suffer a sequence of consistent and complex changes. The NVU is both a structural and conceptual framework that recognizes functional interactions among the cells and their environment, during normal conditions and following injury. This conceptual unit consists of microvessels (endothelial cells-basal lamina matrix-astrocyte end-feet [and pericytes]), astrocytes, neurons and their axons, and other supporting cells (e.g. , microglia and oligodendroglia) that are likely to modulate the function of the unit. This also provides a framework for considering potential bidirectional communication between neurons and their supply microvessels with the participation of intervening astrocytes. The failure of “neuroprotectant” agents, intended to “protect” the neurons during ischemic injury, to bring clinical benefit can be explained by targeting of only one component of the neurovascular unit. On the contrary, maintenance or restitution of blood flow to the unit appears to produce benefit to the NVU. Among the numerous potential interrelationships among cells and components of the neurovascular unit, the contributions of astrocytes and microglial cells to the integrity of the unit during injury have not been rigorously explored. For instance, the now well-described responses of the gelatinases (pro-MMP-2 and pro-MMP-9, and their activated products) have been shown to increase variously in small and large animal models of focal cerebral ischemia, and in various settings in ischemic stroke patients. A rigorous examination of that data suggests inconsistencies among the clinical reports that have been left unexplained, and the exact cellular sources are not named. As the potential window for treatment with large artery recanalization techniques increases the possible ill effects of these proteases on the NVU become more important to understand. A critical examination of the roles of astrocytes and microglia in NVU responses to ischemia can resolve many of these clinical inconsistencies. (pro-) MMP-2 and (pro-) MMP-9 generation in non-human primate tissue with regions of ischemia and plasma leakage, and primary murine microglial cells and astrocytes, have been assayed by immunocytochemistry, zymography, and real-time RT-PCR. Ischemia-related hemorrhage was associated with microglial cell activation in tissue, and the leakage of plasma fibronectin (FN) and vitronectin (VN). In strict serum-depleted primary cultures, pro-MMP-9 was generated exclusively by microglia exposed to VN and FN found in plasma (not in the normoxic CNS) and was enhanced by experimental ischemia (oxygen-glucose deprivation, OGD) . Primary astrocytes generated exclusively proMMP-2, and not pro-MMP-9, which decreased on each matrix substrate during OGD. Microglia-astrocyte contact enhanced pro-MMP-9 generation in a cell-concentration-dependent manner under OGD. Limited studies have indicated that activated microglial cells do not produce sufficient local activity to alter matrix integrity. Hence, asytrocytes alone do not generate pro-MMP-9. It is likely that the resting microglia in brain, upon encountering plasma or blood during focal ischemia, generate MMP-9. But, it is unclear whether either gelatinase is responsible for increased microvascular permeability as has been asserted. This is relevant to the broadening window for the use of the plasminogen activator rt-PA as asserted by ECASS-III, recently published. Here, patients could be safely treated at 3.0-4.5 hours after stroke onset. The safety relative to treatment within a 6.0 hour window in other studies could represent refined patient selection in ECASS-III, to reduce the risk of increased microvessel permeability. This choice suggests that the status of the NVU depends upon patient selection. These observations support a number of implications about how stroke acutely injures both non-vascular and vascular tissues within the brain. They also have implications for the manner in which components of the NVU may com-

Arterial tortuosity syndrome in two Italian paediatric patients

BackgroundArterial tortuosity syndrome (ATS) (OMIM #208050) is a rare autosomal recessive connective tissue disorder characterized by tortuosity and elongation of the large and medium-sized arteries, propensity to aneurysms formation, vascular dissection, and pulmonary arteries stenosis. ATS is caused by mutations in SLC2A10 gene, encoding for the facilitative glucose transporter 10 (GLUT10). So far, 17 SLC2A10 mutations have been reported in 32 families, two of which were Italian with a total of five patients. Here we present the clinical and molecular characterization of two novel Italian paediatric ATS patients.MethodsThe exons and intronic flanking regions of SLC2A10 gene were amplified and direct sequencing was performed.ResultsIn both patients, the involvement of major- and medium-sized arteries was characteristic; the nonvascular connective tissue manifestations were mild and not pathognomic of the disorder. Both patients, born from non-consanguineous parents, were heterozygous for two different SLC2A10 mutations, three of which were recurrent and one was novel (p.Arg231Trp). This mutation is localized at the endofacial loop between the transmembrane domains 6 and 7 of GLUT10.ConclusionTwo novel ATS patients were characterized at clinical and molecular level. Overall, four ATS unrelated families are known in Italy so far. Though ATS clinical delineation improved in the last years, further works in the comprehension of disease presentation and complications onset, particularly in paediatric age, and on ATS molecular basis are needed to add new insights for diagnosis and prevention strategies for related complications.

Use of the Abdominal Rectus Fascia as a Nonvascularized Allograft for Abdominal Wall Closure After Liver, Intestinal, and Multivisceral Transplantation

Abdominal wall closure management has become an important challenge during recipient candidate selection, at the time of donor to recipient matching and during the planning of the surgical procedure for intestinal or multiorgan transplantation. Different strategies have been proposed to overcome the lack of abdominal domain: to reduce the graft size or to increase the abdominal domain. Based on the recent concept of using an acellular dermis matrix (Alloderm) and the availability of abdominal wall tissues from the same organ donor, we conceived the idea of using the fascia of the rectus muscle (FoRM) as a nonvascularized tissue allograft. This is a retrospective report of a series of 16 recipients of FoRM as part of a liver, intestinal, or multiorgan transplant procedure performed between October 2004 and May 2008 at three different transplant centers. Of the 16 recipients of FoRM, all but one case was performed during their transplantation (four multivisceral, two modified multivisceral, three isolated intestine, and two livers). Five patients underwent a retransplant surgery (two livers, two multivisceral, and one isolated intestine). Abdominal wall infection was present in 7 of 16 cases. Nine patients are still alive. No deaths were related to wound infection. Long-term survival showed complete wound healing and only one ventral hernia. The use of a nonvascularized FoRM is a novel and simple surgical option to resolve complex abdominal wall defects in liver/intestinal/multivisceral transplant recipients when it can be covered with the recipient skin.

Cellular xenotransplantation

Expectations are high on cellular therapy. Being fundamental to elucidate organogenesis, it is unlikely that embryonic stem cells will be used for clinical purposes. Postembryonic stage, developing cells are, therefore, the front-runner for regenerative medicine. In addition to autologous cells, both allogeneic and xenogeneic cells are hypothetical candidates to treat specific diseases. This review summarizes the current knowledge on immunological and functional aspects of xeno(allo)-cellular transplantation for cardiomyopathy, diabetes, liver failure, neural diseases, and bone regeneration. Xenocellular transplantation is promising for tissue repair in immunologically privileged sites such as the central nervous system or nonvascularized tissues in which no or moderate immunosuppression is required. In vascularized organs, major immune responses are present when cells are transplanted without additional conditioning. Positive results from encapsulation methods that protect cells from the immune system should further stimulate preclinical research. Also, conditioning immunosuppression could be used to circumvent the initial immune response. Transgenic pigs cells are probably the best xenogeneic substitute for human application, although basic research on innate and noninnate immunity toward pig cells is still required. In several fields of medicine, cellular xenotransplantation is slowly emerging as a potential therapeutic tool.

Rapid estrogen signaling via GPR30 in coronary artery smooth muscle

There is no clear understanding of how coronary artery function is regulated by estrogen (E2), which can either relax or contract arteries via rapid, non‐genomic mechanisms involving classic estrogen receptors (ER). Our objective was to investigate non‐genomic E2 signaling mechanisms in coronary artery smooth muscle. In addition to ERα and ERβ, E2 may also stimulate a G protein‐coupled estrogen receptor, GPR30, in non‐vascular tissue; however, a potential role for GPR30 in coronary arteries is unknown. We found that G1, a selective GPR30 agonist, produces a concentration‐dependent relaxation of endothelium‐denuded porcine coronary arteries in vitro (EC50, 26.5 nM). Further, this response was inhibited by 1mM tetraethylammonium, an inhibitor of large‐conductance calcium‐activated potassium (BKCa) channels. Interestingly, ICI182,780, a non‐selective ER antagonist which can activate GPR30, also induced a concentration‐dependent relaxation of endothelium‐denuded coronary arteries (EC50, 137 nM) contracted with PGF2α, but was far less effective in relaxing arteries contracted with 30mM KCl. Immunoblot studies detected expression of GPR30 protein in smooth muscle from either porcine or human coronary arteries. These studies suggest a novel estrogen signaling mechanism in coronary arteries: stimulation of GPR30, leading to potassium efflux and membrane hyperpolarization. (Supported by HL73890)

Cystathionine γ‐lyase and cystathionine β‐synthase expression and activity change with age and caloric restriction

Hydrogen sulfide (H2S), a gas signaling molecule with actions on the cardiovascular system, is endogenously produced by cystathionine γ‐lyase (CSE) and cystathionine β‐synthase (CBS). We have previously shown an increase in H2S signaling efficacy with age and caloric restriction (CR) in rat aorta and hypothesized that CSE and CBS expression would change similarly. Aorta and liver tissues were collected from rats of ages 8 to 38 months maintained on CR or ad libitum (AL) diets. Protein and RNA were extracted for western blotting and real time PCR, respectively. H2S production was determined in tissue homogenates. While age did not affect H2S production or RNA expression, diet did: aorta and liver H2S production and CSE RNA expression were higher in CR than AL animals for all ages (t‐test, p=0.041). Age and diet both affected protein expression (2‐way ANOVA, p=0.0056): in aorta, both enzymes increased with age, with expression highest in AL animals at older ages; in liver, CSE increased with age, with expression highest in CR animals at older ages, whereas CBS decreased with age with no diet affect. These data show that for aorta, enzyme expression increases with age while RNA expression does not, and CR animals with lower CSE and CBS produce more H2S than AL animals. However, this may not be true of non‐vascular tissues, such as liver. This suggests a decrease in enzyme function with age that may be attenuated by CR.

The C Terminus of the Polerovirus P5 Readthrough Domain Limits Virus Infection to the Phloem

Poleroviruses are restricted to vascular phloem tissues from which they are transmitted by their aphid vectors and are not transmissible mechanically. Phloem limitation has been attributed to the absence of virus proteins either facilitating movement or counteracting plant defense. The polerovirus capsid is composed of two forms of coat protein, the major P3 protein and the minor P3/P5 protein, a translational readthrough of P3. P3/P5 is required for insect transmission and acts in trans to facilitate long-distance virus movement in phloem tissue. Specific potato leafroll virus mutants lacking part or all of the P5 domain moved into and infected nonvascular mesophyll tissue when the source-sink relationship of the plant (Solanum sarrachoides) was altered by pruning, with the progeny virus now being transmissible mechanically. However, in a period of months, a phloem-specific distribution of the virus was reestablished in the absence of aphid transmission. Virus from the new phloem-limited infection showed compensatory mutations that would be expected to restore the production of full-length P3/P5 as well as the loss of mechanical transmissibility. The data support our hypothesis that phloem limitation in poleroviruses presumably does not result from a deficiency in the repertoire of virus genes but rather results from P3/P5 accumulation under selection in the infected plant, with the colateral effect of facilitating transmission by phloem-feeding aphid vectors.

Morphological and molecular aspects of physiological vascular morphogenesis

The cardiovascular system plays a crucial role in vertebrate development and homeostasis. Several genetic and epigenetic mechanisms are involved in the early development of the vascular system. During embryonal life, blood vessels first appear as the result of vasculogenesis, whereas remodeling of the primary vascular plexus occurs by angiogenesis. Many tissue-derived factors are involved in blood vessel formation and evidence is emerging that endothelial cells themselves represent a source of instructive signals to non-vascular tissue cells during organ development. This review article summarizes our knowledge concerning the principal factors involved in the regulation of vascular morphogenesis.

ASYMMETRIC LEAVES2-LIKE19/LATERAL ORGAN BOUNDARIES DOMAIN30andASL20/LBD18Regulate Tracheary Element Differentiation inArabidopsis

ASYMMETRIC LEAVES2 (AS2)/LATERAL ORGAN BOUNDARIES DOMAIN (LBD) family proteins are plant-specific nuclear proteins, and genes encoding several family members have been implicated in plant development. We investigated the function of two members of the Arabidopsis thaliana AS2/LBD family, AS2-LIKE19 (ASL19)/LBD30 and ASL20/LBD18, which encode homologous proteins. Both ASL19 and ASL20 were expressed in immature tracheary elements (TEs), and the expression was dependent on VASCULAR-RELATED NAC-DOMAIN PROTEIN6 (VND6) and VND7, which are transcription factors required for TE differentiation. Overexpression of ASL19 and ASL20 induced transdifferentiation of cells from nonvascular tissues into TE-like cells, similar to those induced upon VND6/7 overexpression. By contrast, aberrant TEs were formed when a cDNA encoding a fusion protein of ASL20 with an artificial repressor domain (ASL20-SRDX) was expressed from its native promoter. These results provide evidence that ASL proteins positively regulate TE differentiation. In transgenic plants overexpressing both ASL19 and ASL20, the xylem-deficient phenotype caused by the expression of dominant-negative versions of VND6/7 proteins was not rescued. However, ectopic expression of VND7 was detected in plants overexpressing ASL20. Moreover, VND genes and their downstream targets were downregulated in ASL20-SRDX plants. Therefore, ASL20 appears to be involved in a positive feedback loop for VND7 expression that regulates TE differentiation-related genes.

Nonvascular contribution to ecosystem NPP in a subarctic heath during early and late growing season

Bryophytes and lichens abound in many arctic ecosystems and can contribute substantially to the ecosystem net primary production (NPP). Because of their growth seasonality and their potential for growth out of the growing season peak, bryophyte and lichen contribution to NPP may be particularly significant when vascular plants are less active and ecosystems act as a source of carbon (C). To clarify these dynamics, nonvascular and vascular aboveground NPP was compared for a subarctic heath during two contrasting periods of the growing season, viz. early-mid summer and late summer-early autumn. Nonvascular NPP was determined by assessing shoot biomass increment of three moss species (Hylocomium splendens, Pleurozium schreberi and Dicranum elongatum) and by scaling to ecosystem level using average standing crop. For D. elongatum, these estimates were compared with production estimates obtained from measurements of shoot length increase. Vascular NPP was determined by harvesting shrub and herb apical growth and considering production due to stem secondary growth of shrubs. Hylocomium splendens and Pleurozium schreberi showed highest biomass growth in late summer, whereas for D. elongatum this occurred in early summer. Maximum relative growth rates were ca. 0.003–0.007 g g−1 d−1. For D. elongatum, production estimates from length growth differed from estimations from biomass growth, likely because of an uncoupling between length growth and biomass shoot growth. Nonvascular NPP was 0.37 and 0.46 g dry weight m−2 d−1, in early and late summer, respectively, whereas in the same periods vascular NPP was 3.6 and 1.1 g dry weight m−2 d−1. The contribution of nonvascular NPP to total aboveground NPP was therefore minor in early summer but substantial in late summer, when 25% of the C accumulated by the vegetation was incorporated into nonvascular plant tissue. The expected global change-induced reduction of nonvascular plant biomass in subarctic heath is likely therefore to enhance C release during the late part of the growing season.

‘Entourage’ effects of N‐palmitoylethanolamide and N‐oleoylethanolamide on vasorelaxation to anandamide occur through TRPV1 receptors

The endocannabinoid N-arachidonoylethanolamide (anandamide) is co-synthesized with other N-acylethanolamides, namely N-palmitoylethanolamide (PEA) and N-oleoylethanolamide (OEA), which have been shown to potentiate anandamide responses (so-called 'entourage effects') in non-vascular tissues. It remains unclear whether such interactions occur in the circulation. In rat isolated small mesenteric arteries, the effects of PEA and OEA on relaxation to anandamide and tissue contents of the N-acylethanolamides were examined under myographic conditions. Anandamide-induced relaxation was potentiated by pretreatment with PEA (10 microM) or OEA (1 microM), or in combination. The potentiation by PEA and OEA was endothelium-independent and abolished by treatment with capsaicin (10 microM), which desensitizes the transient receptor potential vanilloid type 1 (TRPV1) receptor system, or by the TRPV1 receptor antagonist, N-(3-methoxyphenyl)-4-chlorocinnamide (SB366791) (2 microM). It was also observed at molar ratios of anandamide and PEA (or OEA) similar to those found in mesenteric arteries. PEA and inhibition of anandamide hydrolysis by 3'-carbamoyl-biphenyl-3-yl-cyclohexylcarbamate (URB597) (1 microM) additively potentiated anandamide responses. On the other hand, PEA and OEA also induced vasorelaxation per se (rank order of potency: anandamide>OEA>PEA), but relaxation to the three N-acylethanolamides displayed different sensitivity to treatment with capsaicin, SB366791 and URB597. For example, relaxations to anandamide and OEA, but not PEA, were attenuated by both capsaicin and SB366791. This study shows that PEA and OEA potentiate relaxant responses to anandamide through TRPV1 receptors in rat small mesenteric arteries. The congeners also induce vasorelaxation per se, suggesting a function for the N-acylethanolamides in vascular control.

Epoxyeicosatrienoic Acids Function as Selective, Endogenous Antagonists of Native Thromboxane Receptors: Identification of a Novel Mechanism of Vasodilation

Epoxy- and dihydroxy-eicosatrienoic acids (EETs and DHETs) are vasoactive cytochrome P450 metabolites of arachidonic acid. Interestingly, however, the mechanism(s) by which EETs/DHETs mediate smooth muscle relaxation remains unclear. In contrast to previous reports, where dilation was purportedly large-conductance Ca(2+)-activated K(+) (BK(Ca)) and/or transient receptor potential cation channel, subfamily V, member 4 (TRPV4) channel-mediated, 14,15-EET-induced vasodilation [reversal of contractile tone established with the thromboxane receptor (TP) agonist 15-hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic acid (U-46619)] was unaltered in BK(Ca) and TRPV4 knockout mouse isolated aortae compared with wild-type controls, indicating a significant BK(Ca)/TRPV4-resistant mechanism. Whereas all EET and DHET regioisomers reversed U-46619 contraction in rat aortae and mouse mesenteric resistance arteries, these eicosanoids failed to alter phenylephrine-induced contraction, suggesting that they mediated dilation via a "TP-selective" mechanism. Competitive TP antagonism was also observed in nonvascular tissue, including rat fundus and tertiary bronchus, indicating that the effect is not specific to blood vessels. Such effects were TP-selective because 14,15-EET failed to inhibit "non-TP" prostanoid receptor-mediated function in multiple cell/tissue-based assays (K(b) > 10 microM). In accordance, 14,15-EET inhibited specific [(3)H]7-(3-((2-((phenylamino)carbonyl)hydrazino)-methyl)-7-oxabicyclo(2.2.1)hept-2-yl)-5-heptenoic acid (SQ-29548) binding to human recombinant TP receptor, with a K(i) value of 3.2 microM, and it showed weaker affinity for non-TP prostanoid receptors, including DP, FP, EP(1-4), and IP receptors (K(i) values of 6.1, 5.3, 42.6, 19.7, 13.2, 20.2, and >25 microM, respectively) and no appreciable affinity (K(i) values >10 microM) for a diverse array of pharmacologically distinct receptors, including the leukotriene receptors Cys-LT(1/2) and BLT(1). As such, EETs/DHETs represent a unique class of "endogenous" G protein-coupled receptor competitive antagonists, inducing vasodilation via direct TP inhibition. Thus, EETs/DHETs represent novel autoregulatory agents, directly modulating the actions of cyclooxygenase-derived eicosanoids following arachidonic acid mobilization.

Vascular distribution of nitric oxide synthase and vasodilation in the Australian lungfish, Neoceratodus forsteri

The presence of nitric oxide synthase (NOS) and role of nitric oxide (NO) in vascular regulation was investigated in the Australian lungfish, Neoceratodus forsteri. No evidence was found for NOS in the endothelium of large and small blood vessels following processing for NADPH-diaphorase histochemistry. However, both NADPH-diaphorase histochemistry and neural NOS immunohistochemistry demonstrated a sparse network of nitrergic nerves in the dorsal aorta, hepatic artery, and branchial arteries, but there were no nitrergic nerves in small blood vessels in tissues. In contrast, nitrergic nerves were found in non-vascular tissues of the lung, gut and kidney. Dual-wire myography was used to determine if NO signalling occurred in the branchial artery of N. forsteri. Both SNP and SIN-1 had no effect on the pre-constricted branchial artery, but the particulate guanylyl cyclase (GC) activator, C-type natriuretic peptide, always caused vasodilation. Nicotine mediated a dilation that was not inhibited by the soluble GC inhibitor, ODQ, or the NOS inhibitor, L-NNA, but was blocked by the cyclooxygenase inhibitor, indomethacin. These data suggest that NO control of the branchial artery is lacking, but that prostaglandins could be endothelial relaxing factors in the vasculature of lungfish.

Rudimentary leaf abortion with the development of panicle in litchi: Changes in ultrastructure, antioxidant enzymes and phytohormones

Floral bud of litchi ( Litchi chinensis Sonn.) is a mixed bud that consists of leaf primordia, rudimentary leaves and panicle primordia. According to field observation, the fate of rudimentary leaves depends upon the environment and development of the panicle primordia. Vigorous development of panicle results in abortion of rudimentary leaves. In this study, three types of rudimentary leaves, the upper leaves (UL) which subtended a developing panicle primordium and were bound to abscise after a period of panicle development, the abscising lower leaves (AL) which had an emerging axillary panicle and would abscise under a light external force, and the non-abscising lower leaves (NAL) which subtended a vegetative bud and would develop normally, were sampled from the terminal shoots of cvs. ‘Nuomici’ and ‘Feizixiao’. Ultra-structure of the petiole base in the three types of rudimentary leaves in ‘Nuomici’ was observed. Signs of cell autolysis, e.g., tonoplast disruption and fusion of vacuole and cytoplasm, were found in localized cells in UL, while massive cell death was observed in AL, suggesting that the two types of rudimentary leaves were at different stages of abortion. The UL was at an early stage of abortion, whereas AL at a late stage of abortion. Cell autolysis was not observed in the nonvascular tissues in NAL. In ‘Nuomici’, AL had a significantly lower superoxide dismutase (SOD) activity compared to the UL and the NAL, while the latter two had no significant difference in the enzyme activity. In ‘Feizixiao’, however, no significant difference was found among the three types of rudimentary leaves, although AL had the lowest SOD activity. Peroxidase (POD) activity in AL in both cultivars was significantly higher than that in NAL. The UL had a lower POD activity in ‘Nuomici’, but a higher activity in ‘Feizixiao’ compared to NAL. Ascorbate peroxidase (APX) activity was the highest in NAL, and there was no significant difference between the UL and AL. H 2O 2 concentration was the highest in AL in the two cultivars. The NAL had higher levels of zeatin plus zeatin riboside (ZRs), auxin (IAA) and gibberellin (GA 4+7), and lower levels of abscisic acid (ABA) than AL. Total concentration of IAA, GA 4+7 and ZRs (TC) and the ratio of (IAA+GA 4+7 + ZRs)/ABA (RTC) in NAL were higher than those in UL and AL in the two cultivars. The RTC in UL was higher than that in AL. The results suggested that abortion of the rudimentary leaves caused by panicle development may involve programmed cell death, reduced antioxidant activity and increased H 2O 2 and share similar hormonal control (reduced ZRs, IAA and GA 4+7 and increased ABA) to the senescence and abscission of mature leaves.

Clinical Practice Guideline: Penetrating Zone II Neck Trauma

STATEMENT OF THE PROBLEM Penetrating wounds of the neck are common in the civilian trauma population. Risk of significant injury to vital structures in the neck is dependent on the penetrating object. For gunshot wounds, approximately 50% (higher with high velocity weapons) of victims have significant injuries, whereas this risk may be only 10% to 20% with stab wounds. The management of injuries to the neck that penetrate the platysma is dependent on the anatomic level of injury. The neck has been divided into threes zones. Zone I, including the thoracic inlet, up to the level of the cricothyroid membrane, is treated as an upper thoracic injury. Zone III, above the angle of the mandible, is treated as a head injury. Zone II, between zones I and III, is the area of controversy. Because of the density of vital structures in this zone, multiple injuries are common and can affect length of stay. Mortality, particularly for major vascular injuries may reach 50%. Delayed complications such as pseudoaneurysms or arteriovenous fistulae can affect long-term outcomes. Appropriate and timely management of these injuries is critical. For the patients with hard signs of significant injury, including active hemorrhage, expanding hematoma, bruit, pulse deficit, subcutaneous emphysema, hoarseness, stridor, respiratory distress, or hemiparesis, immediate operative management may be indicated. Controversy arises over management of the patient without significant symptoms. The management of these patients has been evolving from an era of mandatory exploration to an era of more selective management. Mandatory exploration, while seemingly safe and conservative, led to many nontherapeutic operations. This fact, along with advances in technology, such as high resolution computed tomography (CT), may eliminate the need to explore the neck to determine whether there are injuries. Also during the time that technology had been advancing, many reports have documented the safety of selective management of neck injuries that penetrate the platysma. This experience has demonstrated that physical examination may be reliable and that not all injuries to vital structures in the neck need surgical intervention for repair. This guideline was therefore initiated to examine the specific roles of mandatory exploration versus selective management based on physical examination and current imaging technologies for penetrating neck trauma.

Epigenetic Regulation of Nr3c1 Expression in Hyperhomocysteinemia

Epigenetic regulation of gene expression may play a role in the endothelial dysfunction associated with hyperhomocysteinemia (HHcy). The objective of this study is to determine if changes in glucocorticoid receptor gene (Nr3c1) expression contributes to HHcy‐related vascular pathology. Nr3c1 expression is regulated by epigenetic mechanisms in non‐vascular tissue and glucocorticoids are vasculoprotective. Mice heterozygous for disruption of the cystathionineβ‐synthase gene (Cbs +/−) and C57BL/6 mice (Cbs +/+) were fed a control diet or a high methionine/low folate (HH) diet to induce HHcy. Total Nr3c1 mRNA levels in aorta, determined by real‐time PCR, were lower (p<0.001) in Cbs +/− and Cbs +/+ fed the HH diet with higher plasma total homocysteine levels than Cbs +/+ mice fed the control diet (22.6±5.7 and 7.17±1.8 vs 2.44±0.2 M, respectively, p<0.001). The 5’ portion of the Nr3c1 gene contains multiple first exons producing heterogeneous RNA transcripts, expressed in a tissue‐specific fashion. We identified expression of two transcripts in aorta arising from exons 1E and 1F and bisulfite pyrosequencing found increased methylation of the exon 1E and exon 1F promoter regions in mice with HHcy. These findings are the first to show methylation‐silencing of vascular Nr3c1 expression and suggest a role for epigenetic regulation of gene expression in HHcy‐related endothelial dysfunction.

Differential regulation of oxidative stress by NO in vascular and nonvascular tissues of endotoxemic rats

Endotoxemic shock is a systemic inflammatory response that is associated with increased NO production by iNOS which contributes to the fall in blood pressure (BP), vascular hyporeactivity and multiple organ failure. NO has also been reported to have both stimulatory and inhibitory effects on oxidative stress (OS). We have previously demonstrated that endotoxin (ET)‐induced fall in BP is associated with an increase in nitrite levels in serum, kidney, heart, thoracic aorta (TA) and superior mesenteric artery (SMA), a decrease in MDA levels in the kidney, heart, TA and SMA, and an increase in MPO activity in the heart and TA, but a decrease in the kidney and SMA of rats. We further investigated the effect of NO on the markers of OS in other tissues of the endotoxemic rats. ET‐induced increase in nitrite production was associated with a decrease in GSH levels in the liver, lungs, brain, spleen and femoral artery (FA). MPO activity was increased by ET in the lungs, spleen and FA, but was decreased in the liver and brain. MDA levels were increased by ET in the lungs, brain, spleen and FA, but were decreased in the liver. SOD and catalase activity was decreased in the liver and spleen, but was increased in the lungs, brain and FA. The effects of ET were prevented by an iNOS inhibitor. The data suggest that NO differentially regulates OS depending on the tissue during endotoxemia. This study was supported by Mersin University Research Foundation.