Immunohistochemistry

Long-term success of distraction osteogenesis depends on the ability of the surrounding soft tissues to tolerate distraction forces and to adapt to the resulting increase in skeletal length and volume. The hypothesis tested in this study was that mandibular elongation by distraction induces myocyte proliferation. Unilateral mandibular angle osteotomies were performed in 16 Yucatan minipigs. The hemimandibles were lengthened using semiburied distraction devices (Synthes Maxillofacial, Paoli, PA) with 0-day latency, 1, 2, or 4 mm/d distraction rates, and a neutral fixation period of twice the gap size in days. In 2 additional animals, the dissection and osteotomies were performed, and distraction devices were placed without activation (sham control). At the end of neutral fixation, tissues were taken from masseter muscle overlying the osteotomy and the equivalent region on the unoperated side (contralateral control). Proliferation of myocytes was estimated using immunohistochemical localization with antibodies against proliferating cell nuclear antigen (PCNA). Muscle overlying the distracted mandible showed 6-fold more PCNA-positive myocytes (16.8% +/- 11.3%) than the contralateral control side (2.8% +/- 1.1%, P < .0001). In the 2 sham-control animals, there was a low index of PCNA-positive myocytes on both the osteotomy (2%, 5%) and the contralateral sides (1%, 2%). The results of this study suggest that distraction of the porcine mandible by the protocol described induces myocyte proliferation in the masseter muscle. A proliferative response may contribute to improved long-term stability of mandibular expansion by distraction osteogenesis.

Our aim was to study histopathological changes in lung tissue at the light microscopic and ultrastructural level during recovery from immunosuppression and Pneumocystis carinii pneumonia. Male Wistar rats were immunosuppressed by per oral dexamethasone for 12 weeks to induce P. carinii pneumonia, after which dexamethasone was stopped. Recovery was monitored 1, 2 and 4 weeks after cessation of the immunosuppression. In immunosuppressed animals, CD4+ and CD8+ lymphocytes were both decreased in situ. CD8+ lymphocytes increased above control level at week one. Like CD8+ cells, the ED1+ macrophages increased rapidly in situ. This was accompanied by a progressively increasing migration (more transient for lymphocytes) of macrophages into bronchoalveolar fluid, associated with morphological signs of activation and phagocytosis and proliferation of type II pneumocytes. Bronchoalveolar lavage (BAL) fluid tumour necrosis factor-alpha (TNF-alpha) increased from subnormal levels to a 4 week pea...

Mitogen- and stress-activated kinases (MSKs) are important substrates of the mitogen-activated protein kinase (MAPK)-activated protein kinase family. MSK1 and MSK2 are both nuclear serine/threonine protein kinases, with MSK1 being suggested to potentially play a role in breast cancer cell proliferation, cell cycle progression, cell migration, invasion and tumour growth. The aim of the current study was to assess MSK1 protein expression in breast cancer tumour specimens, evaluating its prognostic significance. A large cohort of 1902 early stage invasive breast cancer patients was used to explore the expression of MSK1. Protein expression was examined using standard immunohistochemistry on tissue microarrays. Low MSK1 protein expression was associated with younger age (P = 0.004), higher tumour grade (P < 0.001), higher Nottingham Prognostic Index scores (P = 0.007), negative ER (P < 0.001) and PR (P < 0.001) status, and with triple-negative (P < 0.001) and basal-like (P &...

Tissue hypoxia results from an inadequate supply of oxygen (O2) that compromises biological functions. Structural and functional abnormalities of the tumour vasculature together with altered diffusion conditions inside the tumour seem to be the main causes of tumour hypoxia. Evidence from experimental and clinical studies points to a role for tumour hypoxia in tumour propagation, resistance to therapy and malignant

The subcellular localization of five isoforms of facilitated-diffusion glucose transporters (GLUTs), from GLUT1 to GLUT5, in rat pancreatic islets was studied by immunohistochemistry using rabbit polyclonal antisera against mouse or rat GLUT peptides. Animals were perfusion-fixed with phosphate-buffered 4% paraformaldehyde and the pancreases were removed. Some specimens were embedded in paraffin, serially sectioned, and immunostained for glucagon, insulin, somatostatin, and the GLUTs for light microscopic observation. Others were prepared for immunoelectron microscopy by the post-embedding method. By these methods, GLUT2 immunostaining was observed on the lateral membranes of pancreatic beta-cells, whereas GLUT3 immunoreaction was predominantly localized in the cytoplasm of beta-cells and was not found in alpha-cells. In contrast, GLUT5 immunostaining was preferentially localized in the cytoplasm of alpha-cells compared to that of beta-cells. However, GLUT1 and GLUT4 were either barely or not at all detectable in any cells. These results suggest that rat islets take up glucose by at least three different processes and that blood glucose levels could be modulated differentially by: a high Km glucose transporter, GLUT2, in beta-cells; by a low Km glucose transporter, GLUT3, in beta-cells; and by a low Km glucose transporter, GLUT5, in alpha-cells.

Transforming growth factor alpha (TGFα) belongs to the epidermal growth factor (EGF) family and is known to play an important role during eyelid morphogenesis. In this study, we showed that ectopic expression of TGFα in the stroma of Kera-rtTA/tet-O-TGFα bitransgenic mice results in precocious eye opening, abnormal morphogenesis of the meibomian gland, tendon and tarsal plate malformation and epithelium hyperplasia. TGFα did not change proliferation and differentiation of meibocytes, but promoted proliferation and inhibited differentiation of the tarsal plate tenocytes. These results suggest that proper formation of the tendon and tarsal plate in the mouse eyelid is required for normal morphogenesis of the meibomian gland.

Pancreatic cancer is a rapidly fatal disease, and there is an urgent need for early detection markers and novel therapeutic targets. The current study has used a proteomic approach of two-dimensional (2D) gel electrophoresis and mass spectrometry (MS) to identify differentially expressed ...

The aim of this study was to determine the extent of accumulation of lipoprotein(a) [Lp(a)] in human arterial wall and to define its potential role in atherogenesis. Biopsies routinely taken from the ascending aorta of 107 patients undergoing aortocoronary bypass surgery were analyzed for lipid and lipoprotein parameters, which were then correlated to serum values. A significant positive correlation was established between serum Lp(a) and arterial wall apolipoprotein (apo)(a) by enzyme-linked immunosorbent assay. High serum Lp(a) also led to a significant increase of apo B in the arterial wall. No significant correlation was found between apo B in serum and aortic tissue. Apo B was found to be partially linked to apo(a) in the aortic extract. Furthermore, apo(a) was found to be intact, as determined by its molecular weight in sodium dodecyl sulfate electrophoresis. This technique also revealed that the apo(a) isoform pattern of aortic homogenate was comparable to the individual seru...

Parathyroid hormone-related protein (PTHrP) is the major mediator of the humoral hypercalcemia of malignancy and of malignant osteolysis associated with skeletal metastases of common epithelial cancers. PTHrP secretion is regulated by the extracellular calcium concentration ([Ca(2+)](o)) in several types of normal and malignant cells. Because the [Ca(2+)](o)-sensing receptor (CaR) is a key mediator of [Ca(2+)](o)-regulated hormone secretion [e.g., of parathyroid hormone (PTH) by parathyroid chief cells], we investigated the expression of the CaR and PTHrP in normal and neoplastic glial cells and studied the effects of [Ca(2+)](o) on PTHrP secretion. Our results show that primary embryonic human astrocytes (HPA) express CaR mRNA and protein as detected by RT-PCR and Western analysis, respectively. Furthermore, astrocytomas and meningiomas also express the CaR at similar levels as assessed by RT-PCR and Northern and Western blot analyses. HPA and astrocytomas express transcripts encod...

Limb girdle muscular dystrophy type 2A (LGMD2A) is caused by mutations in the calpain 3 gene. In a large family affected by LGMD2A with four severely affected members, three additional asymptomatic relatives had very high serum creatine kinase concentrations. All were homozygous for the R110X mutation and showed a total absence of calpain 3 in the muscle. Histological analysis of

Special AT-rich sequence binding protein 1 (SATB1) is a genome organizer that is expressed by T cells. T cell development is severely impaired in SATB1 null mice; however, because SATB1 null mice die by 3 wk of age, the roles of SATB1 in T cell development have not been well clarified. In this study, we generated and analyzed SATB1 conditional knockout (cKO) mice, in which the SATB1 gene was deleted from all hematopoietic cells. T cell numbers were reduced in these mice, mainly because of a deficiency in positive selection at the CD4(+)CD8(+) double-positive stage during T cell development in the thymus. We also found that SATB1 cKO mice developed autoimmune diseases within 16 wk after birth. In SATB1 cKO mice, the numbers of Foxp3(+) regulatory T (Treg) cells were significantly reduced at 2 wk of age compared with wild-type littermates. Although the numbers gradually increased upon aging, Treg cells in SATB1 cKO mice were still less than those in wild-type littermates at adulthood....

SUMMARY In tetrapods, arginine vasopressin and its counterpart, arginine vasotocin(AVT), are involved in renal water conservation through vascular V1a-type and tubular V2-type receptors, and only the former has thus far been cloned in fish. We successfully cloned the V1a-type and V2-type AVT receptor from the kidney of the African lungfish, Protopterus annectens, and the deduced amino acid sequences exhibited high homology with amphibian V1a- and V2-type receptors, respectively. Functional analysis showed that AVT addition to CHO cells transfected with lungfish V1a-type receptor increased[Ca2+]i in a concentration-dependent manner, whereas CHO cells transfected with lungfish V2-type receptor responded with cAMP accumulation after AVT stimulation. Lungfish V2-type receptor mRNA was strongly expressed in the heart and kidney, while V1a-type receptor mRNA was ubiquitously expressed in all the tissues examined. In the kidney,immunohistochemistry using a specific antibody to lungfish V2-...