Dual role of osteoblastic proenkephalin derived peptides in skeletal tissues

Proenkephalin encodes a group of small peptides with opiate-like activity, the endogenous opioids, known to function as neurohormones, neuromodulators, and neurotransmitters. Recently, we have demonstrated that in addition to its abundance in fetal brain tissue, proenkephalin is highly expressed in nondifferentiated mesodermal cells of developing fetuses. We identified the skeletal tissues, bone, and cartilage as major sites of proenkephalin expression. To examine the possibility that proenkephalin is involved in bone development we have studied the expression of this gene in bone-derived cells, its modulation by bone active hormones, and the effects of enkephalin-derived peptides on osteoblastic phenotype. Our studies revealed that osteoblastic cells synthesize high levels of proenkephalin mRNA which are translated, and the derived peptides are secreted. Reciprocal interrelationships between osteoblast maturation and proenkephalin expression were established. These results together with our observations demonstrating inhibitory effects of proenkephalin-derived peptides on osteoblastic alkaline phosphatase activity, strongly support the notion that proenkephalin is involved in bone development. A different direction of research by other investigators has established the capability of the opioid system in the periphery to participate in the control of pain. On the basis of these two lines of observation, we would like to present the following hypothesis: The potential of embryonic skeletal tissue to synthesize proenkephalin-derived peptides is retained in the adult in small defined undifferentiated cell populations. This potential is realized in certain situations requiring rapid growth, such as remodeling or fracture repair. We suggest that in these processes, similarly to the situation in the embryo, the undifferentiated dividing cells produce the endogenous opioids. In the adult these peptides may have a dual function, namely participating in the control of tissue regeneration and in the control of pain. © 1994 Wiley-Liss, Inc.     

Indoleamine 2,3-Dioxygenase Tissue Distribution and Cellular Localization in Mice: Implications for Its Biological Functions

Earlier studies have suggested that indoleamine 2,3-dioxygenase (IDO) has a wide tissue distribution in mammals. However, detailed information on its cellular localization and also the levels of expression in various tissues is still scarce. In the present study, we sought to determine the cellular localization of IDO and also to quantify the level of its expression in various mouse tissues by using the branched DNA signal amplification assay, Western blotting, and immunohistochemical staining. The highest levels of constitutive IDO expression were found to be selectively present in the caput of epididymis, except for its initial segment. IDO expression was also detected inside the luminal compartment and even in the stereocilia within this region. In the prostate, high levels of IDO were selectively expressed in the capsular cells. In addition, high levels of IDO expression were also selectively detected in certain types of cells in the placenta, spleen, thymus, lung, and digestive tract. Notably, the morphological features of most of the positively stained cells in these organs closely resembled those of antigen-presenting cells. Based on the tissue distribution and cellular localization characteristics of IDO, it is hypothesized that its expression may serve two main functions: one is to deplete tryptophan in an enclosed microenvironment (such as in the epididymal duct lumen) to prevent bacterial or viral infection, and the other is to produce bioactive tryptophan catabolites that would serve to suppress T-cell–mediated immune responses against self-antigens, fetal antigens, or allogeneic antigens, in different situations. (J Histochem Cytochem 58:17–28, 2010)     

Analysis of folate cofactor levels in tissues using high-performance liquid chromatography

A method for the isolation and concentration of the monoglutamate forms of folate cofactors from tissues and for their subsequent separation and quantitation using HPLC coupled with uv detection at 284 nm is described. A chromatographic procedure utilizing Dowex 50 has been developed for the separation of the folate monoglutamates from a large portion of the nonfolate-related material following digestion of the polyglutamated froms with a highly purified preparation of rat liver conjugase. This chromatographic procedure combined with concentration of the Dowex eluate by lyophilization eliminates uv-absorbing material, which interferes with the detection and quantitation of the folate cofactors and makes possible uv measurement of the individual folates. Reverse-phase paired-ion chromatography on μBondapak C₁₈ coupled with uv detection allows direct quantitation of the folates in the nanogram range.     

Six-year field test results of micropropagated black cherry (Prunus serotina)

Summary A field test was established in 1987 to evaluate the growth of micropropagated black cherry plantlets and control seedlings. The study also evaluated effects of two container types on initial survival and growth and of pruning on stem form and growth. At the time of field establishment, plantlets had more extensive root systems than the control seedlings. Survival and height growth were not influenced by container size. Through the first three growing seasons, seedlings were larger than micropropagated plants, but growth differences diminished in the fourth through sixth seasons. Pruning increased the length of clear stem by nearly five-fold but adversely affected diameter growth. Although all clones were from ortet trees more than 50 yr old, none showed plagiotropic growth. Six-year results showed that in a well-prepared and maintained plantation, black cherry trees derived from tissue culture can have at least 80% survival, and growth rates in excess of 1 m per year.     

Flow cytometric analysis of somaclonal variation in lineages of Hosta sports detects polyploidy and aneuploidy chimeras

Somaclonal variation of some 124 specially selected cultivars of Hosta Tratt. (Hostaceae) was investigated. Nuclear DNA contents (2C‐value) were measured by flow cytometry of leaves and roots of L1, L2 and L3 layers derived from apical meristems. These values were then converted to inferred ploidies by comparing the measured 2C‐values and ploidy with those of the parent plant. During tissue‐culture propagation, on occasion diploid (L1‐L2‐L3 = 2‐2‐2) hostas give rise to polyploids, such as fully tetraploids (4‐4‐4), and periclinal chimeras, such as partial tetraploids (4‐2‐2). Continual propagation can result in partial tetraploids becoming full tetraploids. Nuclear DNA of some diploids increased with incomplete chromosome sets resulting in fully aneuploids, such as hostas with a DNA ploidy of L1‐L2‐L3 = 2.5‐2.5‐2.5 and 3.7‐3.7‐3.7, and even in aneuploid periclinal chimeras, such as L1‐L2‐L3 = 2.5‐2‐2 and 3.8‐2‐2. The polyploidy of L1, irrespective of the ploidy of L2 and L3, is found to mainly determine the thickness of leaves. Also the higher the ploidy of L1, the wider and more intense in color is the leaf margin. The measurements of Hosta cultivars and their lineages of sports show that chromosome losses or gains are an important source of new cultivars. The complexity of chromosomal distribution in lineages of several Hosta cultivars is discussed.     

The optimization of a scaffold for cartilage regeneration

Natural polymers offer various advantages in cartilage tissue engineering applications, thanks to their intrinsic bioactivity and adaptability, which can be exploited for the optimization of scaffold properties. In particular, silk fibroin has multifunctional features driven by the self-assembly of molecular subunits in appropriate environmental conditions. For these reasons, it was used in combination with hyaluronic acid to produce porous sponges for cartilage regeneration. The added amount of hyaluronic acid and the cross-linking with genipin modulated scaffold properties in a synergistic way, showing a strong inter-correlation among macroscopic and microscopic characteristics. Interestingly, hyaluronic acid affected silk fibroin conformation and induced a physical separation between the two material components in absence of genipin. Instead, this was prevented by the cross-linking reaction, resulting in a more interspersed network of protein and polysaccharide molecules partially resembling the structure of cartilage extracellular matrix. In addition, the systematic evaluation of sponge properties and how they can be modulated will represent a significant starting point for the interpretation of the complex outcomes driven by the scaffold in vitro and in vivo.     

Peripheral Serotonin: a New Player in Systemic Energy Homeostasis

Whole body energy balance is achieved through the coordinated regulation of energy intake and energy expenditure in various tissues including liver, muscle and adipose tissues. A positive energy imbalance by excessive energy intake or insufficient energy expenditure results in obesity and related metabolic diseases. Although there have been many obesity treatment trials aimed at the reduction of energy intake, these strategies have achieved only limited success because of their associated adverse effects. An ancient neurotransmitter, serotonin is among those traditional pharmacological targets for anti-obesity treatment because it exhibits strong anorectic effect in the brain. However, recent studies suggest the new functions of peripheral serotonin in energy homeostasis ranging from the endocrine regulation by gut-derived serotonin to the autocrine/paracrine regulation by adipocyte-derived serotonin. Here, we discuss the role of serotonin in the regulation of energy homeostasis and introduce peripheral serotonin as a possible target for anti-obesity treatment.