Quantitative analysis of type IV collagen alpha chains in the basement membrane of human urogenital epithelium

Type IV collagen is a major component of the basement membrane (BM), which consists of six genetically distinct (IV) chains. In this study the expression of these six (IV) chains was demonstrated immunohistochemically. In addition, the 2(IV) and 5(IV) chains were analysed quantitatively by confocal laser scanning microscopy in human urogenital epithelial BM. The 1/2(IV) and 5/6(IV) chains were immunoreactive in the epithelial BM, whereas, 3/4(IV) chains were not. The quantitative analysis revealed that the amount of 2(IV) and 5(IV) chains differed in each urogenital epithelial BM. The content of 5(IV) chains in the epithelial BM of the bladder was differentially high, and that of the foreskin was differentially low. It is concluded that the elasticity of epithelial BM of the bladder may be structurally related to the high content of 5/6(IV) chains.     

Immunohistochemical and ultrastructural evidence for myelopoiesis in the scid/scid mouse thymus

The ultrastructure of scid mouse thymus (a small encapsulated epithelial mass within the precardial fat pad) is described. The epithelium did not form cortex or medulla and hence remained relatively undifferentiated. Small unmyelinated nerves innervated the capsule, the major blood vessels and were distributed between the epithelial cells. Fenestrated blood vessels were common. Thymocytes were not identified but numeous granulocytes, mast cells and some fibroblasts, macrophages and interdigitating cells were present. All stages of granulopoiesis were observed in scid thymus. A very small number of immunoreactive ER-MP58 cells indicated bone marrow derived myeloid precursor cells, and low numbers of ER-MP12⁺ and ER-MP20⁺ mononuclear cells indicated stages of myeloid cells committed to the granulocyte/macrophage lineage. Cells containing proliferating nuclear cell antigen (cells in G1, S and G2-M stage) were present throughout the thymic mass. BALB/c thymuses contained cortical foci of p53⁺ cells whereas in scid mice, p53 positive cells were scattered singly throughout the thymus. This study indicates that the presence of moderately extensive myelopoiesis within the scid mouse thymus has potential for the study of extramedullary hematopoiesis, and also is important to bear this function in mind when using the scid mouse as an immunological model for thymus reconstitution and for creating organoid cultures.     

Heterogeneity of epithelial cells in the human thymus

Summary To evaluate interrelationships among epithelial cells, and between morphology and function in the microenvironment, we studied the ultrastructural morphology of epithelial cells in sections of human thymus from donors aged 2 months to 31 years. Six types of epithelial cells were observed: subcapsular-perivascular (type 1); pale (type 2); intermediate (type 3); dark (type 4); undifferentiated (type 5); and large-medullary (type 6). Cells of types 2, 3 and 4 were found throughout the organ. The type-2 to -4 epithelial cells may represent various stages in a differentiation process. In this, type-2 cells are very active and type-4 cells are possibly degenerating elements. Type-4 cells can also contribute to Hassall's corpuscles. Type-5 cells were located mainly in the cortico-medullary region and showed the morphological characteristics of undifferentiated elements. Type-6 cells were located exclusively in the medulla and displayed characteristics of cellular activity. Small Hassall's corpuscles consisted of type-6 epithelial cells; in larger corpuscles many nuclei of type-6 cells were found. Cells of types 2 and 6 contained tubular structures (diameter approximately 20 nm).Concerning the function of thymus epithelial cells, the features associated with protein synthesis observed in cellular types 2 and 6 make them likely candidates for humoral factor-producing and/or secreting elements. In addition, type-2 and -3 cells in the cortex appear to contribute to a special pattern of epithelium-lymphocyte interaction (thymic nurse cells), as demonstrated by the intracytoplasmic location of lymphocytes in the epithelial cells. The various steps in intrathymic T-cell maturation occur at locations in a microenvironment composed of morphologically distinct epithelial cells.     

Multinucleate cells in thymus of C3H mice

Summary Multinucleate epithelial cells occur in the thymus of C3H mice. They are poorly differentiated and scarce, but are more numerous in the medulla than in the cortex. Their increase in number with age is particularly significant between the first and the third months especially for cells with a large number of nuclei, and may be related to thymic involution.Viral particles of type C, similar to those described in murine leukemias, are found in mono- and multinucleate medullary epithelial cells.Research supported by grant 10.013 of the Fonds de la Recherche Fondamentale Collective (Brussels)     

Distribution of the α2-macroglobulin receptor/low density lipoprotein receptor-related protein in human tissues

Summary The hepatic ₁-macroglobulin receptor (₂MR)/low density lipoprotein receptor-related protein (LRP) binds and endocytoses ₂-macroglobulin-proteinase complexes in plasma. In addition, it binds lipoproteins, a novel 40 kDa protein, and complexes between plasminogen activators and plasminogen activator inhibitor type-1. This study shows, for the first time, the tissue distribution of ₂MR/LRP as determined by immunohistochemistry with specific monoclonal antibodies. The analysis revealed ₂MR/LRP-expression in a restricted spectrum of cell types, including neurons and astrocytes in the central nervous system, epithelial cells of the gastrointestinal tract, smooth muscle cells, fibroblasts, Leydig cells in testis, granulosa cells in ovary, and dendritic interstitial cells of kidney. Monocytederived cells displayed marked ₂MR/LRP expression in the phagocytes of liver, lung and lymphoid tissues, but no or low expression in antigen-presenting cells including Langerhans' cells of the skin. The high abundance of ₂MR/LRP in certain cell types of most organs suggests two main routes for ₂MR/LRP ligand clearance: (1) systemic removal in liver of circulating ligands, and (2) non-hepatic interstitial removal in different organs, including the brain.     

Thymic nurse cells: morphological study during their isolation from murine thymus

Summary Thymic nurse cells (TNC), which are multicellular complexes composed of epithelial cells and thymocytes, were obtained from C3H-mice thymuses. They were described by means of light and electron microscopy. The morphology of epithelial cells forming isolated TNC compared to that of small tissue fragments obtained by enzymatic digestion revealed that TNC could be derived from all parts of the thymus: cortex, corticomedullary junction and medulla, the cortex being their principal source. This variety of origin, the presence of several epithelial cells inside a single TNC, the presence of non-lymphoid cells, and the various locations of eleaved desmosomes confirmed that their aspect in vitro as round and sealed structures can be considered to be an artifact due to the isolation technique used. Indeed, during this procedure, they are formed by a process of wrapping of the epithelial cytoplasm around the tightly associated thymocytes. All three epithelial cell types: cortical reticular cells, medullary reticular cells, and medullary globular cells can form TNC.A portion of this work was presented at the first Thymus Workshop. Rolduc, Netherlands, April, 1988     

Immunolocalization of ecto-5′-nucleotidase in the kidney by a monoclonal antibody

Summary A monoclonal antibody IgG, has been raised against ecto-5-nucleotidase purified from rat kidney homogenate. The specificity of the antibody was verified by immunoprecipitation. The distribution of the corresponding antigen in the rat kidney was studied by immunocytochemistry (FITC and PAP technique) in 1 m thick cryostat sections. The antibody reacted with the brush border of proximal tubules, the apical cell membrane and the apical cytoplasm of intercalated cells in connecting tubules and collecting ducts and with interstitial cells of the cortex. Among the interstitial cells exclusively stellate shaped fibroblasts were reactive whereas rounded interstitial cells (type II interstitial cells) as well as pericytes and endothelial cells of peritubular capillaries were unreactive. Compared to the staining intensity of the fibroblasts in the cortical labyrinth the reactivity of the fibroblasts in the medullary rays of the cortex was weak or absent. Interstitial cells of the entire medulla were unreactive. Concerning the fibroblasts in the periarterial connective tissue, those surrounding the larger arteries (arcuate arteries, cortical radial arteries) were negative, those alongside afferent and efferent arterioles were positive. Endothelia of lymphatic capillaries travelling within the periarterial connective tissue were also positive. All components of the juxtaglomerular apparatus were negative.The findings are consistent with an interstitial production of adenosine, available extracellularly and thus being able to reach the major target sites of adenosine, the smooth muscles of glomerular arterioles, including the granular cells at the glomerular vascular pole.     

Immunoreactive neurotensin and somatostatin in the chicken thymus

Summary Two distinct populations of endocrine cells in the chicken thymus display neurotensin and somatostatin immunoreactivity, respectively. Both cell types are few in number at hatching but proliferate rapidly during the first week. The neurotensin cells are Grimelius-positive and Hellerström-Hellmannegative. The somatostatin cells are Grimelius-negative and Hellerström-Hellman-positive. Both cell populations are non-argentaffin. The somatostatin-like material extracted from chicken thymus behaves immunochemically and chromatographically similar to synthetovine somatostatin, while the neurotensin-like material, from the thymus as well as from the gut, differs from synthetic bovine neurotensin in that it appears larger in size and more basic.     

Influence of fibroblasts on epidermization by keratinocytes cultured on synthetic porous membrane (insert) at the air-liquid interface

Culture of keratinocytes on a noncoated porous synthetic membrane maintained at the air-liquid interface allows the establishment of a fibroblast/keratinocyte co-culture, without direct cell-cell contant between the two cellular layers. The influence of fibroblasts (proliferating, confluent or blocked by mitomycin C) on epidermization (i.e., expression of integrins and markers of epidermal differentiation) was studied by immunohistochemistry in two culture media. In the medium supplemented with VCS or Ultroser G and in the absence of fibroblasts, 2, 3, 5 and 6 subunits of integrins are expressed by the basal keratinocytes, except 5 which does not appear with the medium supplemented with Ultroser G. During stratification, the 3 subunit is the only one to persist on suprabasal cells and all the markers of epidermal differentiation studied (filaggrin, involucrin, transglutaminase, keratins K1/K10) are expressed at the 14th day of emerged culture. The presence of fibroblasts modifies the expression profile of integrins: when they are proliferative, the expression of 2 and 6 chains is delayed in the medium supplemented with FCS, and the 6 chain is absent in the medium supplemented with Ultroser G; when they are confluent or blocked by mitomycin C, greater changes are observed only in the medium supplemented with Ultroser G and lead to inhibition or delay of the expression of 2 and 6. In the presence of fibroblasts, only the expression of filaggrin (marker of terminal differentiation) is affected; it is delayed in the medium supplemented with FCS whatever the state of fibroblasts, and is inhibited in the medium supplemented with Ultroser G in the presence of proliferating and confluent fibroblasts.Abbreviations DMEM Dulbecco's Modified Eagle's Medium - EGF epidermal growth factor - K-SMM keratinocyte-serum free medium - PBS phosphate-buffered saline Ca²⁺- and Mg²-free     

Formation ofα,ω-dodecanedioic acid andα,ω-tridecanedioic acid from different substrates by immobilized cells of a mutant ofCandida tropicalis

Summary The metabolic formation of either,-dodecanedioic acid or,-tridecanedioic acid from the individual n-alkane, n-alcohol, n-monoacid and,-diol with corresponding carbon chain length using K-carrageenan entrapped mutants S76 ofCandida tropicalis was studied. The immobilized cells of S76 could also directly produce-hydroxy acid and,-dioic acid from,-diol. With n-alcohol and n-monoacid as substrate, the amount of-hydroxy acid and,-dioic acid produced was also a function of the incubation time.The results demonstrated that in the immobilized cells of S76 the formation of,-dioic acid from n-alcohol can also run both via n-monoacid and via,-diol as well as in the normal cells of S76.     

Cyclic nucleotide phosphodiesterases of the renal cortex

Summary Cyclic nucleotide phosphodiesterase in the basal-lateral segment of plasma membranes from proximal tubule cells of the rabbit renal cortex was studied and compared to that in the brush border segment of the plasma membrane. Both adenosine 3,5-monophosphate and guanosine 3,5-monophosphate were hydrolyzed by the basal-lateral membrane, but activity varied differently with the two substrates in a complex concentration-dependent manner. Activity with adenosine 3,5-monophosphate was greater than, equal to, or less than with guanosine 3,5-monophosphate, at concentrations of 1000, 100, and 10 to 1 m, respectively. Basal-lateral membrane phosphodiesterase activities at 1 and 500 m substrate exhibited differential responses to pH, metals, heat, and a heat stable inhibitor. Stimulation by guanosine 3,5-monophosphate and inosine 3,5-monophosphate of adenosine 3,5-monophosphate hydrolysis was found in basal-lateral but not in brush border membranes. This stimulation was potentiated by ethyleneglycol-bis(-aminoethyl ether)N,N-tetraacetic acid and ethylenediaminetetraacetate, inhibited by Triton X-100, and totally blocked by Zn²⁺. The findings indicate that multiple forms of phosphodiesterase are present in the basal-lateral segment and these differ from the activities in the brush border region of the plasma membrane. The characteristics of (i) allosteric, guanosine 3,5-monophosphate-sensitivity of adensoine 3,5-monophosphate phosphodiesterase, and (ii) relatively high guanosine 3,5-monophosphate phosphodiesterase activity, in basal-lateral membranes, which are also enriched in adenylate and guanylate cyclase, suggest an important physiological role for these phosphodiesterases in the regulation of net production of cyclic nucleotides in the renal cortex.     

Immunohistochemical localization of human α2macroglobulin in connective tissue

Summary The localization of ₂ macroglobulin (₂M) has been examined by an indirect immunofluorescent technique in frozen sections of various human tissues. The results indicate that ₂M is present only in connective tissues and blood. The outer medulla of the kidney and the submucosa of the gut showed the strongest reaction. Epithelia or endothelial cells were unreactive. In liver, only the Kupffer cells were stained. These results were confirmed with the immunoperoxidase technique and by the study of tissue extracts in crossed immunoelectrophoresis (CIE). As a positive control a polyspecific antiserum prepared against whole human fibroblasts as well as anti-albumin were used. Our findings are interpreted in the light of the observations that ₂M is synthesized and selectively ingested by fibroblasts.Abbreviation used in this Paper ₂M ₂macroglobulin This work was supported by a Grant from the Belgian Cancer Fund (Algemene Spaar en Lijfrente Kas), by Grant 3.0043.79 (Fonds voor Geneeskundig Wetenschappelijk Onderzoek) and by Research Fund OT/VII/30 (K.U. Leuven)F. Van Leuven is a Post Doctoral Research Fellow of the American Cystic Fibrosis Foundation     

Ecto-5′-nucleotidase is expressed by pericytes and fibroblasts in the rat heart

Ecto-5-nucleotidase is anchored at the outer surface of cell membranes and thus its reaction product adenosine is released into the extracellular space. Extracellular adenosine displays via specific receptors a wide range of physiological effects in heart. There are discrepancies in the literature concerning the distribution of ecto-5-nucleotidase in heart. Since we suspected that these may be due to technical problems, in the present study on ecto-5-nucleotidase in rat heart we attempted to circumvent some technical pitfalls. Good preservation of the tissue with open capillary lumina, providing a clear identification of endothelium, was obtained by perfusion fixation. At the light microscopic level, the distribution of ecto-5-nucleotidase studied by enzyme histochemistry and immunohistochemistry using a monoclonal and a polyclonal antibody yielded congruent results. The enzyme was rather homogeneously distributed throughout the myocardium, with a slightly higher incidence of stained cells in the outer thirds than in the inner third of the wall. Consistently high levels of ecto-5-nucleotidase were seen only in interstitial cells. The walls of large vessels and heart muscle cells were constantly negative for ecto-5-nucleotidase. The endothelia of capillaries were mostly negative but a few profiles occasionally displayed a weak immunoreaction. The interstitial cells staining positive for ecto-5-nucleotidase could be identified as pericytes and as fibroblasts according to their shapes and localizations. The immunoreactivity of fibroblasts was confirmed by electron microscopy. These data indicate that adenosine may be formed extracellularly in the interstitium of the myocardium, where it would have direct access to important targets such as myocytes, arterioles and nerve endings.     

Patterns of direct and indirect embryogenesis from mesophyll protoplasts of Medicago sativa

Clones of three cultivars of Medicago sativa (Rambler, Regen S and Rangelander) were used as sources of mesophyll protoplasts. Although all three clones readily produced protoplasts, the subsequent development patterns in culture varied greatly among genotypes, with protoplasts from Regen S and Rambler forming calli which could be induced to form embryos, and protoplasts from Rangelander undergoing direct embryogenesis. Protoplasts of Regen S exhibited high rates of division while those of Rangelander tended to aggregate with only a few cells per aggregate surviving. The surviving cells gave rise to proembryos within the aggregates; these proembryos developed into differentiated embryos after 5–7 weeks of culture. Based on the initial protoplast population, the efficiency of embryo formation averaged 0.13% and ranged from 0.001–0.4%. Observations during the early stages of culture indicated that cell aggregation was a prerequisite for direct embryogenesis.     

Interaction of poly(ADP-ribose)polymerase with DNA polymerase α

Homogeneously purified poly(ADP-ribose) polymerase (PARP) specifically stimulated the activity of immunoaffinity-purified calf or human DNA polymerase by about 6 to 60-fold. Apparently, poly(ADP-ribosyl)ation of DNA polymerase was not necessary for the stimulation. The effects of PARP on DNA polymerase were biphasic: at very low concentrations of DNA, it rather inhibited its activity, whereas, at higher DNA concentrations, PARP greatly stimulated it. The autopoly(ADP-ribosyl)ation of PARP suppressed both its stimulatory and inhibitory effects. By immunoprecipitation with an anti-DNA polymerase antibody, it was clearly shown that PARP may be physically associated with DNA polymerase . Stimulation of DNA polymerase may be attributed to the physical association between the two, rather than to the DNA-binding capacity of PARP, since the PARP fragment containing only the DNA binding domain showed little stimulatory activity. The existence of PARP-DNA polymerase complexes were also detected in crude extracts of calf thymus.     

Measurement of H2'-C2' and H3'-C3' dipolar couplings in RNA molecules

The quality of nucleic acid solution structures can be significantly improved using residual dipolar coupling data. However, many of the one-bond couplings that could be used for this purpose are difficult to measure. Conventional 2D experiments are often unable to reveal one-bond H2-C2 and H3-C3 couplings in large RNA molecules due to spectral overlap. Here we show how to use 3D HCcH-COSY and Relay HCcH-COSY to measure one-bond H2-C2 and H3-C3 couplings which improved the precision of the structures obtained recently for a 42 nucleotide RNA.     

High D-glucose does not affect binding of α-tocopherol to human erythrocytes

The role of -tocopherol uptake system in human erythrocyte in the uptake of plasma -tocopherol has been suggested. However no information is available on -tocopherol uptake activity of human erythrocytes in the presence of high levels of D-glucose which is known to lead to pathological alterations in different cells including human erythrocytes. Therefore, in order to examine the effect of D-glucose on the binding of -tocopherol to human erythrocytes, the binding characteristics of -tocopherol to these cells were established first. Binding of [3H]-tocopherol to human erythrocytes was both saturable and specific. Scatchard analysis of -tocopherol binding to these cells showed the presence of two independent classes of binding sites with widely different affinities. The high affinity binding sites had a dissociation constant (Kd1) of 90 nM with a binding capacity (n1) of 900 sites per cell, whereas the low affinity binding sites had a dissociation constant (Kd2) of 5.2 M and a binding capacity (n2) of 105,400 sites per cell. Trypsin treatment abolished all the -tocopherol binding activity. Competition for the binding of -tocopherol to human erythrocytes was effective with other homologues of -tocopherol (-tocopherol, -tocopherol and -tocopherol) and their potency was almost equal to -tocopherol itself. The order of preference was -tocopherol > -tocopherol -tocopherol -tocopherol. Incubation of human erythrocytes with various concentrations of D-glucose did not affect -tocopherol uptake activity. Our data demonstrate the presence of an -tocopherol uptake system in human erythrocytes and that the -tocopherol uptake activity is not modulated by the presence of D-glucose.     

Induction of lymphokine-activated killer activity in mice by prothymosin α

We have recently demonstrated that prothymosin (ProT) when administered intraperitoneally (i.p.) protects DBA/2 mice against the growth of syngeneic leukemic L1210 cells through the induction of tumoricidal peritoneal cells producing high levels of tumor necrosis factor (TNF) [Papanastasiou et al. (1992) Cancer Immunol Immunother 35: 145]. In this report we tested further immunological alterations that may be caused by the administration of ProT in vivo. We demonstrate that i.p. injections of ProT enhance natural killer (NK) cell activity and induce lymphokine-activated (LAK) activity in vivo. Thus, splenocytes from ProT-treated DBA/2 animals exhibited significantly higher cytotoxic activity (up to threefold) against the NK-sensitive YAC cell line and the NK-resistant P815 and L1210 syngeneic tumor cells, as compared to splenocytes from syngeneic control mice. The enhancement of the cytotoxic profile of DBA/2 splenocytes was associated with increased percentages of CD8⁺ cells, NK cells and activated CD3⁺ cells. The ProT-induced effect persisted for 30 days after the end of the ProT treatment period and returned to normal levels 20 days later. SPlenocytes from non-treated DBA/2 animals generated high NK and LAK activities in response to ProT in vitro. The ProT-induced NK an LAK activities reached 84% and 75% respectively of what was obtained with interleukin-2 (IL-2). High concentrations of TNF and IL-2 were generated in response to ProT in LAK cultures. These findings suggest that ProT may provide an overall protective effect against tumor growth in vivo through induction of NK and LAK activities possibly indirectly via the production of IL-2 and TNF in the spleen, peritoneal cavity and probably other lymphoid organs.This work was supported by a CEC grant to M. Papamichail     

Protonmotive force in Brevibacterium flavum: the lack of intracellular pH homeostasis

Brevibacterium flavum 22LD-P cells were shown to maintain a transmembrane pH gradient (pH) from 0.6 to 1.8–2 units and a transmembrane electric potential difference () from 0 to 200 mV depending on the pH and ionic composition of the incubation medium, grwoth substrate and concentration of cells. decreased from 120–140 mV to 0 when medium pH was lowered from neutral to 5.0–5.5 and increased to 180–200 mV when medium pH was raised to 8–9 in cells utilizing acetate or endogenous substrate. Cells growing on sucrose, kept around 100–120 mV at neutral as well as acidic medium pH. Intracellular pH in the acetate utilizing or endogenously respiring cells was maintained with the range of 8.9 to 5.5 at medium pH ranging from 9.1 to 4.0, respectively. Sucrose grown cells were able to maintain a more stable intracellular pH. Endogenously respiring cells in potassium phosphate buffer at high biomass concentrations maintained larger pH and relatively smaller , than the same cells in diluted suspensions. Cells in sodium phosphate buffer possessed larger and almost no pH, but was still dependent on biomass concentration.The lack of intracellular pH homeostasis and the collapse of at acid medium pH are discussed in the context of cell membrane proton permeability.     

Poly(U)-directed peptide-bond formation from the 2′(3′)-glycyl esters of adenosine derivatives

Summary The self-condensation of 2(3)-O-glycyl esters of adenosine, adenosine-5-(O-methylphosphate) and P₁, P₂-diadenosine-5-pyrophosphate in 6.2 mM solutions at pH 8.0 and -5°C in the presence of 12.5 mM poly(U) yields approximately 3 times as much diketopiperazine as reactions without poly(U). As the concentration of 2(3)-O-(glycyl)-P₁, P₂-diadenosine-5-pyrophosphate is decreased from 6.2 mM to 1.5 mM the yield of diketopiperazine in the presence of poly(U) decreases slightly from 6.6% to 5.2%, whereas, in the absence of poly(U) the yield of diketopiperazine decreases substantially from 2.4% to 0.75%. The enhanced yield of diketopiperazine that is attributed to the template action of poly(U) is temperature dependent and is observed only at temperatures below 10°C (5°C to -5°C) for 6.2 mM 2(3)-O-(glycyl)-adenosine-5-(O-methylphosphate) and below 23°C (15°C to -5°C) for 6.2 mM 2(3)-O-(glycyl)-P₁, P₂-diadenosine-5-pyrophosphate. The absence of a template effect at high temperatures is attributed to the melting of the organized helices. The hydrolysis half-lives at pH 8.0 and -5°C of 2(3)-O-(glycyl)-adenosine, 2(3)-O-(glycyl)-adenosine-5-(O-methylphosphate), 2(3)-O-(glycyl)-P₁, P₂-diadenosine-5-pyrophosphate, and 5-O-(glycyl)-adenosine in the presence of poly(U) are substantially larger than their half-lives in the absence of poly(U). The condensation of 2(3)-O-(glycyl)-adenosine yields 5% of 5-O-(glycyl)-adenosine in the presence of poly(U) compared to 0.7% in the absence of poly(U).Abbreviations DKP diketopiperazine - (gly)₂ glycylglycine - (gly)₃ glycylglycylglycine - AppA-gly 2(3)-O-(glycyl)-P₁, P₂-diadenosine-5-pyrophosphate - MepA-gly 2(3)-O-(glycyl)-adenosine-5-(O-methylphosphate) - Ado-2(3)-gly 2(3)-O-(glycyl)-adenosine - Ado-5-gly 5-O-(glycyl)-adenosine - Boc-gly N-tert-butyloxycarbonylglycine - AppA P₁, P₂-diadenosine-5-pyrophosphate - MepA adenosine-5-(O-methylphosphate) - AppA-Boc-gly 2(3)-O-(Boc-glycyl)-P₁, P₂-diadenosine-5-pyrophosphate - Ado-5-Boc-gly 5-O-(Boc-glycyl)-adenosine - Ado-2(3)-Boc-gly 2(3)-O-(Boc-glycyl)-adenosine