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Aim. The subject of this study is evaluation of histological hallmarks of kidney growth in rats after a surgery and subsequent management of the wound with fibrin glue. Material and Methods. Sixty-five Wistar rats underwent a surgical procedure resulting in parenchymal injury by incision followed by an application of fibrin glue. Kidneys have been examined and evaluated histologically after 4 weeks (group I) and 26 weeks (group II) after the surgery Results. When fibrin glue was used, in both groups, regular anastomoses of the wound edges of the kidney after surgery were noted. Histological features of kidney growth (renal glomeruli, renal tubules and blood vessels) were observed in the parenchyma and in the postoperative scar. In the second group kidneys grew on average by 3.5mm in the longitudinal section and by 1.89 in the cross section. In this group, the number of renal glomeruli nearly doubled. Glomeruli and tubules were also found in the postoperative scar.

The actual incidence of renal dysfunction after contrast media administration seems to be underestimated, especially in the context of epidemiological data. There are only few data concerning the monitoring of impaired kidney function within a few hours after iodine contrast medium application. Hence, the purpose of this study is to observe the incidence of early renal function deterioration within 12-18 h after administration of iodine contrast media in patients scheduled for elective coronary angiography, who were intravenously and orally hydrated. In addition, the project aims to reclassify the contrast induced nephropathy phenomenon, by identification of early markers of renal dysfunction. Morphology, electrolytes, blood urea nitrogen (BUN), creatinine, low-density lipoprotein cholesterol, triglycerides, high-density lipoprotein, and total cholesterol levels were assessed with the use of typical laboratory techniques in 319 patients referred for coronary angiography. We demonstrated that early deterioration of renal function in patients 12-18 h after administration of contrast during imaging tests (even when appropriate prophylactic hydration was used), may occurred just as an increase (or no change) of serum creatinine level and BUN level and a decrease of creatinine clearance and glomerular filtration rate. Depending on the parameter, the phenomenon can be found in 13-28 % of all respondents. Early renal function impairment defined as above was almost 2 and 2.22 × 10(3) times (respectively) more frequently observed in our study than contrast induced nephropathy defined by current definitions.

Background: Qa-2 is a nonclassical MHC Ib antigen, which has been implicated in both innate and adaptive immune responses, as well as embryonic development. Qa-2 has an unusual peptide binding specificity in that it requires two dominant C-terminal anchor residues and is capable of associating with a substantially more diverse array of peptide sequences than other nonclassical MHC. Results: We have determined the crystal structure, to 2.3 A, of the Q9 gene of murine Qa-2 complexed with a self-peptide derived from the L19 ribosomal protein, which is abundant in the pool of peptides eluted from the Q9 groove. The 9 amino acid peptide is bound high in a shallow, hydrophobic binding groove of Q9, which is missing a C pocket. The peptide makes few specific contacts and exhibits extremely poor shape complementarity to the MHC groove, which facilitates the presentation of a degenerate array of sequences. The L19 peptide is in a centrally bulged conformation that is stabilized by intramolecular interactions from the invariant P7 histidine anchor residue and by a hydrophobic core of preferred secondary anchor residues that have minimal interaction with the MHC. Conclusions: Unexpectedly, the preferred secondary peptide residues that exhibit tenuous contact with Q9 contribute significantly to the overall stability of the peptide-MHC complex. The structure of this complex implies a "conformational" selection by Q9 for peptide residues that optimally stabilize the large bulge rather than making intimate contact with the MHC pockets.

Introduction There are scarce data about the effect of metformin on lipid profile in patients with type 1 diabetes. Objectives The present study is the first prospective clinical trial evaluating the effect of combined therapy of metformin and insulin on the pool of oxidized and glycated low‑density lipoproteins (LDL) in young patients with type 1 diabetes and concomitant obesity. Patients and methods A total of 33 obese patients with type 1 diabetes treated with intensive insulin therapy were randomized into a group where metformin was added. The remaining 19 patients continued to receive intensive insulin therapy (control group). In all patients, lipid profile and glycemia were assessed using routine laboratory tests. Oxidized and glycated LDL were measured using commercially available kits. Laboratory tests were performed at baseline and at a control visit after 6 months of treatment. Results A significant decrease in the levels of glycated hemoglobin, fasting plasma glucose, postprandial glucose, average glucose, triglycerides, glycated LDL, and body mass index was observed in the group receiving combined therapy. A similar decrease was not observed in the control group. The remaining lipid parameters were not changed during follow‑up in any of the groups. Conclusions Addition of metformin to intensive insulin therapy in young obese patients with type 1 diabetes results in a significant reduction of glycated LDL levels. This can be possibly explained by better glucose control, which improved insulin sensitivity of the peripheral tissues and reduced body mass in this patient group.

Class Ib Qa-2 molecules are expressed in tissue culture cells as approximately 40-kDa membrane-bound, glycophosphatidylinositol-linked antigens and as approximately 39-kDa soluble polypeptides. Recently, alternative splicing events which delete exon 5 from a portion of Qa-2 transcripts were demonstrated to give rise to truncated secreted Qa-2 molecules in transfected cell lines. To determine whether this mechanism operates in vivo and to find out whether Qa-2 can be detected in soluble form in circulation, murine blood samples were analyzed. Critical to these experiments was preparation of an anti-peptide antiserum against an epitope encoded by a junction of exon 4 and exon 6. We find that supernatants of splenocytes cultured in vitro as well as serum or plasma contain two forms of soluble Qa-2 molecules. One form corresponds to a secreted molecule translated from transcripts from which exon 5 has been deleted; the other is derived from membrane-bound antigens or their precursors. The levels of both soluble forms of Qa-2 are inducible upon stimulation of the immune system, suggesting an immunoregulatory role for these molecules or for the mechanism leading to the reduction of cell-associated Qa-2 antigens in vivo.

Unlike the highly polymorphic major histocompatibility complex (MHC) class Ia molecules, which present a wide variety of peptides to T cells, it is generally assumed that the nonpolymorphic MHC class Ib molecules may have evolved to function as highly specialized receptors for the presentation of structurally unique peptides. However, a thorough biochemical analysis of one class Ib molecule, the soluble isoform of Qa-2 antigen (H-2SQ7b), has revealed that it binds a diverse array of structurally similar peptides derived from intracellular proteins in much the same manner as the classical antigen-presenting molecules. Specifically, we find that SQ7b molecules are heterodimers of heavy and light chains complexed with nonameric peptides in a 1:1:1 ratio. These peptides contain a conserved hydrophobic residue at the COOH terminus and a combination of one or more conserved residue(s) at P7 (histidine), P2 (glutamine/leucine), and/or P3 (leucine/asparagine) as anchors for binding SQ7b. 2 of 18 sequenced peptides matched cytosolic proteins (cofilin and L19 ribosomal protein), suggesting an intracellular source of the SQ7b ligands. Minimal estimates of the peptide repertoire revealed that at least 200 different naturally processed self-peptides can bind SQ7b molecules. Since Qa-2 molecules associate with a diverse array of peptides, we suggest that they function as effective presenting molecules of endogenously synthesized proteins like the class Ia molecules.

Our laboratory has developed a process for generating mAbs with selectivity to unique peptides in the context of MHC molecules. Recently, we reported that RL4B, an mAb that we have called a TCR mimic (TCRm) because it recognizes peptide in the context of MHC, has cytotoxic activity in vitro and prevented growth of tumor cells in a prophylactic setting. When presented in the context of HLA-A2, RL4B TCRm recognizes the peptide GVLPALPQV derived from human chorionic gonadotropin (hCG)-β. In this study, we show that RL4B TCRm has strong binding affinity for the GVLPALPQV peptide/HLA-A2 epitope and fine binding specificity for cells that express endogenous hCGβ Ag and HLA-A2. In addition, suppression of tumor growth with RL4B TCRm was observed in orthotopic models for breast cancer. Using two aggressive human tumor cell lines, MDA-MB-231 and MCF-7, we provide evidence that RL4B TCRm significantly retards tumor growth, supporting a possible role for TCRm agents in therapeutic settings. Moreover, tumors in mice responded to RL4B TCRm therapy in a dose-dependent manner, eliminating tumors at the highest dose. RL4B TCRm strongly detects the hCGβ peptide/HLA-A2 epitope in human primary breast tumor tissue, but does not react or reacts weakly with normal breast tissue from the same patient. These results further illustrate the selective nature of TCRm Abs and the clinical relevance of the GVLPALPQV peptide/HLA-A2 epitope expression in tumor cells, because they provide the first evidence that Abs that mimic the TCR can be used to markedly reduce and suppress tumor growth.

Essential thrombocythemia (ET) co-existing with chronic lymphocytic leukemia (CLL) is extremely rare. We report two cases of ET with Jak 2 V617F in Zap-70+ CLL. ET is a myeloproliferative stem cell disease. Zap-70 expression in CLL correlates with non-mutated immunoglobulin genes. The occurrence of a less mature CLL in patients with a pluripotential stem cell disease raises the possibility that an initial “trigger hit” occurred in a pre-Jak 2 common early progenitor in these patients. Subsequent additional molecular events accumulated independently following myeloid and lymphoid differentiation, leading to the development of two diseases of likely identical origin but different lineages.

Myeloproliferative diseases (MPD) are a recognized risk factor for thrombotic events. Patients with MPD can sometimes present with thrombotic complications even prior to the diagnosis or manifestation of any clinical features of MPD, although the exact incidence of such a mode of presentation of MPD remains unclear, since it had been difficult to make a diagnosis of early or occult MPD without the use of in vitro bone marrow colony assays not commonly available in the routine laboratory. The recent identification of an association between the JAK2 V617F mutation and MPD enables the convenient early diagnosis in these patients using a molecular approach. JAK2 V617F is a gain-of-function mutation that results in a constitutive activation of the JAK-STAT signaling pathway and leads to the expansion of the affected hematopoietic precursor cells, with skewing towards the erythroid lineage. JAK2 V617F mutation has been found in more than 90% of patients with polycythemia rubra vera but with a lesser frequency in the other MPD. Recently, a strong association has been found between JAK2 V617F mutation and idiopathic Budd-Chiari syndrome or splanchnic thrombosis. A study that involved 139 cases of non-cirrhotic splanchnic vein thrombosis identified the JAK2 V617F mutation in 27 (21%) of these patients, 14 of whom had overt MDS, 3 developed MDS during follow up (median 41 months) and 11 were still disease free at the time of reporting (1). Similar studies have been carried out in other common thrombotic diseases but no correlation was found between JAK2 V617F mutation and deep venous thrombosis, pulmonary embolus, stroke or early myocardial infarction (2). It has, therefore, been proposed that the JAK2 V617F mutation should be tested in patients with non-cirrhotic Budd-Chiari syndrome and thrombosis of the splanchnic circulation, even in the absence of any clinical features of MPD. The routine use of this test for other forms of thromboembolic diseases, however, remains debatable. Although the association between splanchnic thrombosis and early MPD is well recognized, the occurrence of arterial stent thrombosis in patients with occult MPD has not previously been reported. We report in this paper the first case of a patient with an occult MPD unmasked by a bilateral early renal arterial stent thrombosis. The diagnosis of an occult MPD was made possible molecularly by the detection of the JAK2 V617F mutation. A 72 years old lady with a history of hypertension and renal impairment was found to have a bilateral renal artery stenosis by CT scan. Her right renal artery was 80–85% and her left renal artery 85–90% stenosed. She underwent a percutaneous bilateral renal arterial angioplasty with insertion of the Dynalink nitinol 6-Fr stents. The success of the procedure was confirmed by a repeat CT scan that showed 100% patency of the stents. The procedure also resulted in an improved renal function. Ten days after the procedure, the patient presented with abdominal and loin pain. Repeat CT scan showed total occlusion of the renal arterial stents by thrombus and the development of renal infarction. The unfortunate and unusual early adverse thrombotic event triggered the investigations for a thrombophilic tendency. A careful examination did not reveal any established genetic risk factors. She did not have any family history of thromboembolic disease. The genetic workup was unremarkable, without any evidence for Factor V Leiden mutation, Prothrombin gene mutation or methyltetrahydrofolate reductase gene mutation. Her serum homocysteine level was not elevated. Serum Protein C, Protein S and antithrombin III were all within limits. Her only risk factor for thromboembolic disease was a history of smoking half a pack of cigarette per day. Antiphospholipid antibody screening was also negative. She had six previous pregnancies without any thromboembolic complications. Her peripheral blood was checked for the JAK2 V617F mutation by PCR because she had a mild leukocytosis. The white cell count of 11 was associated with a normal hemoglobin and platelet count, although her white cell count was normal just before insertion of the renal arterial stents. She was found to be strongly positive for the JAK2 V617F mutation by qualitative PCR on two separate occasions. With further follow-up 3 months, her leukocytosis has resolved spontaneously and she has continued to have a normal complete blood count without developing any clinical features of MPD. Early thrombosis of the arterial stents (

Several new HLA-B (B8, B51, Bw62)- and HLA-C (Cw6, Cw7)-specific genes were isolated either as genomic cosmid or cDNA clones to study the diversity of HLA antigens. The allele specificities were identified by sequence analysis in comparison with published HLA-B and -C sequences, by transfection experiments, and Southern and northern blot analysis using oligonucleotide probes. Comparison of the classical HLA-A, -B, and -C sequences reveals that allele-specific substitutions seem to be rare events. HLA-B51 codes only for one allele-specific residue: arginine at position 81 located on the alpha 1 helix, pointing toward the antigen binding site. HLA-B8 contains an acidic substitution in amino acid position 9 on the first central beta sheet which might affect antigen binding capacity, perhaps in combination with the rare replacement at position 67 (F) on the alpha 1 helix. HLA-B8 shows greatest homology to HLA-Bw42, -Bw41, -B7, and -Bw60 antigens, all of which lack the conserved restriction sites Pst I at position 180 and Sac I at position 131. Both sites associated with amino acid replacements seem to be genetic markers of an evolutionary split of the HLA-B alleles, which is also observed in the leader sequences. HLA-Cw7 shows 98% sequence identity to the JY328 gene. In general, the HLA-C alleles display lower levels of variability in the highly polymorphic regions of the alpha 1 and alpha 2 domains, and have more distinct patterns of locus-specific residues in the transmembrane and cytoplasmic domains. Thus we propose a more recent origin for the HLA-C locus.

The biological properties of the nonclassical class I MHC molecules secreted into blood and tissue fluids are not currently understood. To address this issue, we studied the murine Q10 molecule, one of the most abundant, soluble class Ib molecules. Mass spectrometry analyses of hybrid Q10 polypeptides revealed that α1α2 domains of Q10 associate with 8–9 long peptides similar to the classical class I MHC ligands. Several of the sequenced peptides matched intracellularly synthesized murine proteins. This finding and the observation that the Q10 hybrid assembly is TAP2-dependent supports the notion that Q10 groove is loaded by the classical class I Ag presentation pathway. Peptides eluted from Q10 displayed a binding motif typical of H-2K, D, and L ligands. They carried conserved residues at P2 (Gly), P6 (Leu), and Pω (Phe/Leu). The role of these residues as anchors/auxiliary anchors was confirmed by Ala substitution experiments. The Q10 peptide repertoire was heterogeneous, with 75% of the groove occupied by a multitude of diverse peptides; however, 25% of the molecules bound a single peptide identical to a region of a TCR V β-chain. Since this peptide did not display enhanced binding affinity for Q10 nor does its origin and sequence suggest that it is functionally significant, we propose that the nonclassical class I groove of Q10 resembles H-2K, D, and L grooves more than the highly specialized clefts of nonclassical class I Ags such as Qa-1, HLA-E, and M3.