CO2 back-diffusion in the rete aids O2 secretion in the swimbladder of the eel

In order to estimate the relative importance of back-diffusion of O2 and CO2 for their partial pressure enhancement in the swimbladder, we have determined O2 and CO2 partial pressure and content and pH in microsamples collected non-obstructively from the afferent and efferent rete vessels in the European eel. 1. The PO2 increased significantly along the arterial vessels of the rete (from 33 to 303 Torr), with no change in O2 content, suggesting O2 not to be exchanged in the rete counter-current system. 2. A corresponding increase of PCO2 (from 4 to 35 Torr) was accompanied by a significant rise in CO2 content (from 8 to 15 mmol.L-1), suggesting significant CO2 back-diffusion in the rete. 3. Changes in PO2 during passage of blood through the swimbladder epithelium were variable and small, and the PO2 in rete venous blood was similar to that in rete arterial blood, explaining the lack of O2 back-diffusion. 4. Using blood CO2 dissociation data, about 70% of the rise in arterial CO2 content was estimated to derive from diffusion of CO2, the remaining 30% from diffusion of HCO3-, from venous to arterial rete capillaries, or from H+ transport in the opposite direction. The data indicate that CO2 back-diffusion in the rete does not only raise the rete arterial PCO2; it also reduces the O2 capacity (Root effect) and thus enhances the arterial PO2.     

醛糖还原酶基因5’端(AC)n多态性对2型糖尿病患者红细胞醛糖还原酶活性的影响

目的　探讨醛糖还原酶 (AR)基因 5’端 (AC) n 的多态性对 2型糖尿病 (DM )红细胞AR活性的影响。方法　 16 3例 2型DM分为无微血管病变 (NDC)组 (6 6例 )和微血管病变 (DMAP)组(97例 ) ,正常对照 (CON)组 42例 ;另按AR基因 5’端 (AC) n 的等位基因类型分为DM携带Z 2等位基因 (DZ 2 )组 (5 4例 )、DM携带Z - 2等位基因 (DZ - 2 )组 (35例 )、DM同时携带Z 2和Z - 2等位基因 (Z 2 /Z - 2 )组 (18例 )、DM不携带Z 2和Z - 2等位基因 (X/X)组 (5 6例 )、对照者携带Z 2等位基因 (NZ 2 )组 (2 1例 )和对照者携带Z - 2等位基因 (NZ - 2 )组 (7例 )。用改良Sriratava法测定AR活性并比较其在各组间的差异。结果　DMAP组、NDC组和CON组间的AR活性 (ARA)差异有显著性 ,DMAP组最高 ,NDC组次之 ,CON组最低 (P <0 .0 0 1)。DM组携带Z - 2和Z 2等位基因各亚组中 ,DZ - 2组ARA最高 ,Z - 2 /Z 2和X/X组居中 ,DZ 2组最低 ,差异有显著性统计学意义(P <0 .0 0 1)。DZ - 2和NZ - 2组的ARA分别高于DZ 2和NZ 2组 ,DZ - 2和DZ 2组的ARA分别高于NZ - 2和NZ 2组 (P均 <0 .0 0 1)。结论　AR的激活对DMAP的发生和发展起重要作用。Z - 2等位基因可能是AR的激活因子 ,Z 2等位基因则为其抑制因子。     

Orchidectomy increases the formation of prostanoids and modulates their role in the acetylcholine-induced relaxation in the rat aorta

AIMS: This study examines the effect of endogenous male sex hormones on thromboxane A2 (TXA2), prostaglandin (PG) I2, PGF(2 alpha), and PGE I2 release, as well as their role in acetylcholine (ACh)-mediated relaxation in the aorta. METHODS AND RESULTS: Aortic segments from orchidectomized and control male Sprague-Dawley rats were used to measure COX-2 protein expression. ACh-induced relaxation of these segments was also determined in the absence and presence of the COX-2 inhibitor NS-398, the TXA2 synthesis inhibitor furegrelate, the PGI2 synthesis inhibitor tranylcypromine (TCP), or the thromboxane-prostanoid (TP) receptor antagonist SQ-29 548. Furthermore, TXA2, PGI2, PGF(2 alpha), and PGE2 release as well as the vasomotor effect of exogenous TXA2, PGI2, PGF(2 alpha), and PGE2 were measured. COX-2 expression was increased in aortas from orchidectomized rats. NS-398 did not modify the ACh-induced relaxation in arteries from both control or orchidectomized rats. Furegrelate did not modify the ACh-induced relaxation in aortas from control animals but, in aortas from orchidectomized rats, it increased that response. TCP decreased the ACh-induced relaxation in both groups. The TP receptor antagonist, SQ29 548 failed to modify ACh-induced relaxation in aortas from either rat group. Pre-incubating arteries from orchidectomized rats with TCP plus furegrelate did not modify the decrease in the ACh response induced by TCP alone, but this response was restored by co-incubation of TCP plus SQ29 548. ACh-induced TXA2, PGI2, PGF(2 alpha), and PGE2 release were increased by orchidectomy. The presence of furegrelate plus TCP increased the ACh-induced PGE2 release more in arteries from orchidectomized than in those from control rats. The contractile responses induced by the TXA2 mimetic U-46619 or by exogenous PGF(2 alpha) were similar in arteries from control and orchidectomized rats, while those induced by exogenous PGE2 were increased in arteries from orchidectomized rats; the vasodilator response induced by exogenous PGI2 was decreased in arteries from orchidectomized rats. CONCLUSION: These data show that endogenous male sex hormone deprivation increases COX-2 expression, the release of TXA2, PGI2, PGF(2 alpha), and PGE2 and the contractile response induced by exogenous PGE2 and TXA2, while it decreases the relaxation induced by exogenous PGI2. Despite the predominance of vasoconstrictor prostanoids derived from COX-2 in aortas from orchidectomized rats, the ACh-induced relaxation remains increased.     

IVUS radiofrequency analysis in the evaluation of the polymeric struts of the bioabsorbable everolimus‐eluting device during the bioabsorption process

Background : In the ABSORB study cohort A the changes in the amount of dense calcium and necrotic core have not been reported in comparison to the prestenting phase; this evaluation could be useful to better clarify the bioabsorption process. Aim of this study was therefore to evaluate the dynamic changes in plaque size and plaque tissue composition observed between 6 months and 2 years follow‐up, and to compare these findings to the prestenting phase. Methods : Angiography, intravascular ultrasound and derived parameters (virtual histology, palpography, and echogenicity) were serially assessed postprocedure, at 6 months and at 2 years in 20 patients. In a subset of 8 patients the same measurements were also recorded in the prestenting phase. Results : In the total population a reduction of 18% in the plaque area was observed between 6 month and 2 year follow‐up (7.56 ± 2.32 mm² at 6 months vs. 6.16 ± 2.10 mm² at 2 year follow‐up; P < 0.01). In the subgroup of eight patients who underwent IVUS during the pre‐stenting phase, the plaque area at 2 year follow‐up was not significantly different when compared to the prestenting plaque area (7.29 ± 2.29 mm² at prestenting vs. 7.48 ± 1.45 mm² at 2 year follow‐up, P = NS). Necrotic core area was reduced by 24% between the 6 month and 2 year follow‐up (0.97 ± 0.66 mm² at 6 months vs. 0.74 ± 0.53 mm² at 2 year follow‐up; P = NS), whilst dense calcium was reduced by 14% from 6 month to 2 year follow‐up (0.83 ± 0.50 mm² at 6 months vs 0.72 ± 0.64 mm² at 2 year follow‐up; P = NS). Whilst the necrotic core at 2 years follow‐up was not significantly different when compared to the pre‐stenting phase (0.62 ± 0.42 mm² prestenting vs 1.07 ± 0.56 mm² at 2 year follow‐up; P = NS), the area of dense calcium was significantly higher at follow‐up compared to prestenting (0.35 ± 0.35 mm² pre‐stenting vs. 0.84 ± 0.66 mm² at 2 year follow‐up; P < 0.05). Conclusions : The reduction in the necrotic core component between 6 month and two year follow‐up could be related to a synergistic effect of the bio‐absorption process and the anti‐inflammatory action of everolimus. © 2010 Wiley‐Liss, Inc.     

Molecular basis of the interaction between the antiapoptotic Bcl-2 family proteins and the proapoptotic protein ASPP2

We have characterized the molecular basis of the interaction between ASPP2 and Bcl-2, which are key proteins in the apoptotic pathway. The C-terminal ankyrin repeats and SH3 domain of ASPP2 (ASPP2_Ank-SH3) mediate its interactions with the antiapoptotic protein Bcl-2. We used biophysical and computational methods to identify the interaction sites of Bcl-2 and its homologues with ASPP2. Using peptide array screening, we found that ASPP2_Ank-SH3 binds two homologous sites in all three Bcl proteins tested: (i) the conserved BH4 motif, and (ii) a binding site for proapoptotic regulators. Quantitative binding studies revealed that binding of ASPP2_Ank-SH3 to the Bcl-2 family members is selective at two levels: (i) interaction with Bcl-2-derived peptides is the tightest compared to peptides from the other family members, and (ii) within Bcl-2, binding of ASPP2_Ank-SH3 to the BH4 domain is tightest. Sequence alignment of the ASPP2-binding peptides combined with binding studies of mutated peptides revealed that two nonconserved positions where only Bcl-2 contains positively charged residues account for its tighter binding. The experimental binding results served as a basis for docking analysis, by which we modeled the complexes of ASPP2_Ank-SH3 with the full-length Bcl proteins. Using peptide arrays and quantitative binding studies, we found that Bcl-2 binds three loops in ASPP2_Ank-SH3 with similar affinity, in agreement with our predicted model. Based on our results, we propose a mechanism in which ASPP2 induces apoptosis by inhibiting functional sites of the antiapoptotic Bcl-2 proteins.     

Angiotensin-converting enzyme II in the heart and the kidney

The renin-angiotensin system (RAS) has been recognized for many years as critical pathway for blood pressure control and kidney functions. Although most of the well-known cardiovascular and renal effects of RAS are attributed to angiotensin-converting enzyme (ACE), much less is known about the function of ACE2. Experiments using genetically modified mice and inhibitor studies have shown that ACE2 counterbalances the functions of ACE and that the balance between these two proteases determines local and systemic levels of RAS peptides such as angiotensin II and angiotensin1-7. Ace2 mutant mice exhibit progressive impairment of heart contractility at advanced ages, a phenotype that can be reverted by loss of ACE, suggesting that these enzymes directly control heart function. Moreover, ACE2 is also found to be upregulated in failing hearts. In the kidney, ACE2 protein levels are significantly decreased in hypertensive rats, suggesting a negative regulatory role of ACE2 in blood pressure control. Moreover, ACE2 expression is downregulated in the kidneys of diabetic and pregnant rats and ACE2 mutant mice develop late onset glomerulonephritis resembling diabetic nephropathy. Importantly, ACE2 not only controls angiotensin II levels but functions as a protease on additional molecular targets that could contribute to the observed in vivo phenotypes of ACE2 mutant mice. Thus, ACE2 seems to be a molecule that has protective roles in heart and kidney. The development of drugs that could activate ACE2 function would allow extending our treatment options in diabetic nephropathy, heart failure, or hypertension.     

Cooperative interactions between the interleukin 2 receptor alpha and beta chains alter the interleukin 2-binding affinity of the receptor subunits.

The interleukin 2 (IL-2) receptor (IL-2R) is a multisubunit receptor that includes three major IL-2 binding subunits, the IL-2R alpha, beta, and gamma chains. We have detected and analyzed cooperative interactions between the IL-2R alpha and beta chains (IL-2R alpha and IL-2R beta, respectively) in COS cells transfected with cDNAs encoding the IL-2R alpha, the IL-2R beta, or both cDNAs. We demonstrated that IL-2 F42A, an analog that fails to bind to the isolated IL-2R alpha subunit and would be predicted by the hierarchical affinity-conversion model to have impaired binding to cells expressing both chains, instead readily binds to the IL-2R alpha/beta heterodimer in COS cells. Furthermore, this binding is abolished by the antibody HIEI that separates the two IL-2R subunits. The monoclonal antibodies anti-Tac and Mik-beta 1 directed at the IL-2-binding sites on IL-2R alpha and IL-2R beta, respectively, block ligand binding to the heterodimer. This binding pattern is inconsistent with the strict hierarchical affinity-conversion model that mandates an initial binding of IL-2 to IL-2R alpha followed by binding of the IL-2/IL-2R alpha complex to IL-2R beta. Instead, our results support an alternative model of preformed complexes of IL-2R beta with other IL-2R subunits. In this alternative model, IL-2R alpha and -beta exist in part as preformed complexes in which the affinity of IL-2R beta for IL-2 is altered by the proximity of IL-2R alpha, through mechanisms that do not require the prior binding of IL-2 to IL-2R alpha.     

Bcl-2 decreases the free Ca2+ concentration within the endoplasmic reticulum

The antiapoptotic protein Bcl-2 localizes not only to mitochondria but also to the endoplasmic reticulum (ER). However, the function of Bcl-2 at the level of the ER is poorly understood. In this study, we have investigated the effects of Bcl-2 expression on Ca(2+) storage and release by the ER. The expression of Bcl-2 decreased the amount of Ca(2+) that could be released from intracellular stores, regardless of the mode of store depletion, the cell type, or the species from which Bcl-2 was derived. Bcl-2 also decreased cellular Ca(2+) store content in the presence of mitochondrial inhibitors, suggesting that its effects were not mediated through mitochondrial Ca(2+) uptake. Direct measurements with ER-targeted Ca(2+)-sensitive fluorescent "cameleon" proteins revealed that Bcl-2 decreased the free Ca(2+) concentration within the lumen of the ER, [Ca(2+)](ER). Analysis of the kinetics of Ca(2+) store depletion in response to the Ca(2+)-ATPase inhibitor thapsigargin revealed that Bcl-2 increased the permeability of the ER membrane. These results suggest that Bcl-2 decreases the free Ca(2+) concentration within the ER lumen by increasing the Ca(2+) permeability of the ER membrane. The increased ER Ca(2+) permeability conferred by Bcl-2 would be compatible with an ion channel function of Bcl-2 at the level of the ER membrane.     

Alpha-melanotropin: the minimal active sequence in the lizard skin bioassay

alpha-Melanotropin (alpha-melanocyte-stimulating hormone, alpha-MSH) is a tridecapeptide, Ac-Ser-Tyr-Ser-Met-Glu-His-Phe-Arg-Trp-Gly-Lys-Pro-Val-NH2. The minimal sequence of alpha-MSH required for agonism in the lizard (Anolis carolinensis) skin bioassay was determined to be Ac-His-Phe-Arg-Trp-NH2 (Ac-alpha-MSH6-9-NH2). Smaller fragments of this sequence (Ac-alpha-MSH6-8-NH2, Ac-alpha-MSH6-7-NH2, Ac-alpha-MSH7-9-NH2, and Ac-alpha-MSH7-8-NH2) were devoid of melanotropic activity. The tetrapeptide, Ac-alpha-MSH7-10-NH2, was also inactive, thus again demonstrating the importance of His at position 6 for minimal activity. The important potentiating amino acids were found to be Met-4, Lys-11, and Pro-12, since Ac-alpha-MSH4-10-NH2 was about 100 times more potent than Ac-alpha-MSH5-10-NH2, and Ac-[Nle4]-alpha-MSH4-11-NH2 was about 40 times more potent than Ac-alpha-MSH4-10-NH2 or Ac-[Nle4]-alpha-MSH4-10-NH2. Ac-alpha-MSH4-12-NH2 and Ac-[Nle4]-alpha-MSH4-12-NH2 were equipotent and about six times more potent than alpha-MSH. Since [Nle4]-alpha-MSH and Ac-[Nle4]-alpha-MSH4-13-NH2 were both equipotent but about sixfold less active than Ac-[Nle4]-alpha-MSH4-12-NH2, it is clear that valine at position 13 does not contribute to the potency of alpha-MSH, except possibly in a negative way. The minimal message sequence for equipotency to alpha-MSH appears to be Ac-Met-Glu-His-Phe-Arg-Trp-Gly-Lys-NH2, since the analog, Ac-[Nle4]-alpha-MSH4-11-NH2, was as active as the native hormone. Ser-1, Tyr-2, Ser-3, Glu-5, and Val-13 are not important for melanotropic potency since Ac-alpha-MSH4-12-NH2 was more potent than alpha-MSH, and Ac-alpha-MSH5-10-NH2 and Ac-alpha-MSH6-10-NH2 were equipotent, being about 4,000 times less active than alpha-MSH.     

Oxygen consumption of the chick embryo's respiratory organ, the chorioallantoic membrane

A new technique based on stopping the chick embryo's blood circulation in the intact egg was used to measure in situ the chorioallantoic (CA) oxygen consumption, MCAO2, from incubation day 12 to 20. Total egg MO2, MTOTO2, and wet and dry masses of embryo and CA were also measured daily. Embryo MO2, MEMBO2, was calculated. Mean MCAO2 decreased from 71 mumol X h-1 (17% of MTOTO2, 24% of MEMBO2) at 12 days to 62 mumol X h-1 (5% of MTOTO2) at 20 days. Dry mass of CA did not change significantly. Water remained at a high level in CA (88-94%), but embryo water decreased from 93% to 82% between days 12 and 20. The fairly high level of MCAO2, more marked at young stages, calls for corrections in respiratory and circulatory embryonic variables derived from MTOTO2, such as CA blood flow, CA diffusive capacity for O2, and CA arterio-venous shunt. Mass specific values and intra-specific allometric relations in bird embryos should be recalculated on the basis of MEMBO2 instead of MTOTO2.     

Implications of the Immune Polymorphisms of the Host and the Genetic Variability of SARS-CoV-2 in the Development of COVID-19

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is responsible for the current pandemic affecting almost all countries in the world. SARS-CoV-2 is the agent responsible for coronavirus disease 19 (COVID-19), which has claimed millions of lives around the world. In most patients, SARS-CoV-2 infection does not cause clinical signs. However, some infected people develop symptoms, which include loss of smell or taste, fever, dry cough, headache, severe pneumonia, as well as coagulation disorders. The aim of this work is to report genetic factors of SARS-CoV-2 and host-associated to severe COVID-19, placing special emphasis on the viral entry and molecules of the immune system involved with viral infection. Besides this, we analyze SARS-CoV-2 variants and their structural characteristics related to the binding to polymorphic angiotensin-converting enzyme type 2 (ACE2). Additionally, we also review other polymorphisms as well as some epigenetic factors involved in the immunopathogenesis of COVID-19. These factors and viral variability could explain the increment of infection rate and/or in the development of severe COVID-19.     

Replacement of helium by argon in the measurement of the carbon monoxide diffusing capacity of the lung

Carbon monoxide diffusing capacity of the lung (DLco) of 118 normal subjects and 48 patients was measured by using three different gas mixtures respectively: (1) 10% He, 0.3% CO, 21% O2 and the remainder N2; (2) 10% Ar, 0.3% CO, 21% O2 and the remainder N2; (3) 10% He, 10% Ar, 0.3% CO, 21% O2 and the remainder N2. The results were extremely close with each other, value was 0.99 (P less than 0.0005). These results indicate for the first time that the much cheaper and easy-available gas-argon can be used instead of the very expensive helium for measurement of DLco.     

Nephrotoxicity of the constituents of the gentamicin complex

Commercial gentamicin C is a mixture of gentamicin C1, C1a, and C2. The nephrotoxicity of each of these constituents was compared with that of the gentamicin complex. After seven days the mean creatinine level in serum was 0.8 mg/dl in rats given C2 and 0.5 mg/dl in rats given C1, C1a, or the gentamicin complex (P less than .001). Toxicity attributable to C1a was not detected until day 14, and only minimal toxicity was noted in C1-treated rats after 21 days. Nephrotoxicity caused by the gentamicin complex was similar to that caused by C2. By a new high-pressure liquid chromatographic method, the renal concentration of C1, C1a, and C2 was quantified in rats given the gentamicin complex. The results indicated an early, preferential renal accumulation of C2. Subsequently, the C2 content of 12 commercial lots of gentamicin C was measured. The C2 concentration ranged from 12.4 to 20.1 mg/ml. In short, experimental nephrotoxicity from gentamicin C is largely the result of the C2 constituent, and the concentration of this constituent in commercial preparations of gentamicin varies by as much as 7.7 mg/dl.     

Comparative studies on the metabolism of oestradiol in the brain, the pituitary and the liver of the rat

Radioactive oestradiol was incubated with homogenates of hypothalamus, pituitary and liver and slices of cortex, hippocampus, hypothalamus, pituitary and liver of ovariectomized rats. 2- and 4-substituted oestrogens were the main metabolites in the brain and were found in the same proportions. In the liver only the 2-substituted oestrogens were the main metabolites, while the 4-substituted ones were of minor quantitative importance. On the basis of wet weight the 2-hydroxylating capacity of the liver was approximately 50 fold greater than that of the brain and the 4-hydroxylating capacity about the same as that of the brain. Only slight differences in the 2- and 4-hydroxylating capacity of various brain areas were observed, although apparently the hypothalamus and the pituitary were more active in the formation of catecholestrogens than the cortex and the hippocampus.     

Analysis of the envelope region of the highly divergent HIV-2ALT isolate extends the known range of variability within the primate immunodeficiency viruses.

HIV-2ALT is a highly divergent HIV-2-related isolate that is genetically equidistant to the prototypic HIV-2 strains, defined by HIV-2ROD, and to the simian immunodeficiency viruses SIVmac and SIVsm. We have now cloned and sequenced the envelope region of HIV-2ALT, thus completing the analysis of the whole viral genome. The sequences of env and nef and of the second exons of tat and rev were compared with those of the other viruses of the HIV-2/SIVsm/SIVmac group. Despite of the high degree of variation of HIV-2ALT, functional domains of the genes are conserved. Although in env, the overall pattern of constant and variable domains is maintained, many single amino acid exchanges exist at positions previously thought to be constant in HIV-2 strains. In addition, when compared with a broader spectrum of immunodeficiency viruses, which includes SIVMND from mandrill and SIVAGM from African green monkey, HIV-2ALT Env has a high percentage of amino acid exchanges, which are unique to this strain. This underlines the separate branch of HIV-2ALT within the phylogenetic tree and makes obvious the inclusion of such divergent strains in preventive and therapeutic programs.     

Indomethacin inhibits the effects of oestrogen in the anterior pituitary gland of the rat

Two inhibitors of prostaglandin synthesis, indomethacin and aspirin, blocked the increase of oestrogen-binding sites in the nuclear subcellular fraction, an increase which occurs after the administration of oestradiol. Consequently the biological effects of oestrogens in the anterior pituitary gland of the rat (prolactin synthesis, concentration of progesterone-binding sites and cell proliferation) are diminished. The anterior pituitary gland synthesized prostaglandin F2 alpha (PGF2 alpha), PGE2 and PGD2 from arachidonic acid. This synthesis was blocked when indomethacin was added to the culture media. Oestrogen increased the concentration of PGE2: an increase that was partially prevented by indomethacin. Prostaglandins may have an important role on the effects of oestrogen in the anterior pituitary gland of the rat.     

The influence of the progression of secondary hyperparathyroidism on the set point of the parathyroid hormone-calcium curve

The influence of secondary hyperparathyroidism (2 HPT) on the set point of the parathyroid hormone (PTH)-Ca(2+) curve is controversial. In vitro experiments have shown an increase in the set point. However, clinical studies with hemodialysis patients have provided a variety of results (increases, decreases and no changes in the set point have been reported). The present study was designed to investigate the influence of the progression of 2 HPT on the set point of the PTH-Ca(2+) curve. The PTH-Ca(2+) curve and the expression of parathyroid calcium receptor (CaR mRNA) and vitamin D receptor (VDR mRNA) have been studied in normal rabbits (group I, n=9) and in nephrectomized rabbits (group II, n=18) at two stages after inducing 2 HPT: 2-3 weeks (group IIA) and 5-6 weeks (group IIB). In group I, the set point of the PTH-Ca(2+) curve was 1.63+/-0.03 mM. A progressive hypocalcemia was detected during the evolution of 2 HPT (groups IIA and IIB). Rabbits from group IIA had a significant (P<0.001) decrease in the set point to values of 1.45+/-0.02 mM. However, the set point increased significantly in group IIB (P<0.001) to 1.56+/-0.03 mM. CaR mRNA was similarly decreased in groups IIA (39+/-12%) and IIB (48+/-7%). No changes were detected in VDR mRNA. In conclusion, a reduction in the set point of the PTH-Ca(2+) curve in response to decreased extracellular Ca(2+) was detected in the early phases of 2 HPT. However, with the progression of 2 HPT the set point tended to increase even though extracellular Ca(2+) was markedly decreased. The increase in the set point in the course of 2 HPT seems to be a complex process that cannot be fully explained by changes in parathyroid CaR mRNA or VDR mRNA.