Histology and histopathology Vol.30, nº1 (2015)
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- PublicationOpen AccessThe effect of inhaled nitric oxide on the carrageenan-induced paw edema(F. Hernández y Juan F. Madrid. Universidad de Murcia: Departamento de Biología Celular e Histología, 2015) Coelho, Carly Faria; Vieira, Rodolfo P.; Lopes-Martins, Patrícia Sardinha Leonardo; Teixeira, Simone Aparecida; Borbely, Alexandre Urban; Gouvea, Irene María; Frigo, Lucio; Lopes-Martins, Rodrigo Álvaro BrandãoInhaled nitric oxide therapy reaches not only pulmonary vessels, but also other vasculatures, presenting anti-inflammatory effects. Therefore, this study investigated the effects of inhaled nitric oxide on a mice model of carrageenan-induced paw edema. Paw edema was induced in male Swiss mice (20-30 g) by subplantar injection of carrageenan (0.05 ml of a 1% suspension in 0.9% saline). The evaluation of timecourse edema (mililiter) was measured by plethysmometry until 12 h following carrageenan administration. Thirty minutes after carrageenan injection, some groups received inhaled nitric oxide (300 ppm at variable doses and times) or Indometacin (INDO 5 mg/Kg, v.o), while others received sildenafil (1 mg/Kg, i.p) or rolipram (3 mg/Kg, i.p.) with or without inhaled nitric oxide. Paws were assessed for edema levels by plethysmometry, mieloperoxidase activity and histological analysis. Inhaled nitric oxide significantly reduced carrageenan-induced paw edema, mieloperoxidase activity and inflammatory infiltrate, although similar results were also observed in sildenafil and rolipram treated groups. In addition, significant effects between inhaled nitric oxide with pharmacological therapy was observed. Inhaled nitric oxide presents anti-inflammatory effects on carrageenaninduce paw edema, as observed through reduced edema, mieloperoxidase activity and neutrophil infiltration, indicating that inhaled nitric oxide therapy goes beyond lung vascular effects.
- PublicationOpen AccessThe effect of immunosuppressive therapy on renal cell apoptosis in native rat kidneys(F. Hernández y Juan F. Madrid. Universidad de Murcia: Departamento de Biología Celular e Histología, 2015) Kędzierska, Karolina; Sporniak-Tutak, Katarzyna; Kolasa, Agnieszka; Domański, Leszek; Domański, Maciej; Sindrewicz, Krzysztof; Smektała, Tomasz; Bober, Joanna; Safranow, Krzysztof; Osękowska, Bogumiła; Kabat-Koperska, Joanna; Baranowska-Bosiacka, Irena; Parafiniuk, Mirosław; Urasińska, Elżbieta; Ciechanowski, KazimierzAim: To analyse the impact of the most commonly used immunosuppressive drugs on the occurrence of apoptosis in the native kidneys of Wistar rats. Method: The study involved 36 rats. The animals grouped according to the immunosuppressive regimen used (tacrolimus, mycophenolate mofetil, cyclosporine A, rapamycin and prednisone). The rats in all study groups were treated with a 3-drug protocol for 6 months. The medication dose was adjusted based on available literature data. No drugs were administered to the control group. The rats were then killed. Autopsies of all animals were performed and the kidneys were isolated for histopathology (HE + PAS). To assess cell apoptosis the TUNEL reaction was performed. Blood trough levels of immunosuppressive drugs as well as the parameters of peripheral blood were determined. Results: 1. In rats treated with cyclosporine A distal nephron tubules were characterised by more pronounced apoptosis. 2. In tacrolimus-treated rats a lower intensity of apoptosis was found in the distal tubules. 3. In rapamycin-treated rats the apoptosis was inhibited both in the distal and proximal nephron tubules. 4. In MMF treated rats intense apoptosis was observed in the proximal nephron tubules. 5. There were no significant changes in renal histopathology (HE + PAS). Conclusions: The apoptosis in nephron tubules caused by immunosuppressive therapy is not accompanied by any histopathological changes (eg fibrosis, inflammation, tubular atrophy, vacuolation of the tubular cells) in light microscopy
- PublicationOpen AccessOverexpression of proCOL11A1 as a stromal marker of breast cancer(F. Hernández y Juan F. Madrid. Universidad de Murcia: Departamento de Biología Celular e Histología, 2015) Fuentes Martínez, Nelson; García Pravia, Carmen; García Ocaña, Marcos; Menéndez Rodríguez, Primitiva; Del Amo, Jokin; Suárez Fernandez, Laura; Galván, José A.; De los Toyos, Juan R.; Barneo, LuisBackground: Our previous studies demonstrated the expression of procollagen11A1 in fibroblasts of pancreatic cancer desmoplasia and the lack of expression in fibroblasts of pancreatitis by means of the polyclonal antibody (anti-proCOL11A1 pAb) we generated. In a similar way, we decided to compare the expression of procollagen11A1 in fibroblasts of infiltrating ductal carcinoma of the breast and fibroblasts of benign sclerosing lesions of the breast, in order to validate the anti-proCOL11A1 pAb in this setting and to study how proCOL11A1 expression relates to other prognostic and predictive factors, as well as to survival. Methods: 45 core biopsies of sclerosing adenosis and 50 core biopsies of infiltrating ductal carcinoma of the breast were stained with anti-proCOL11A1 pAb, a polyclonal antibody highly specific to the less homologous fraction of proCOL11A1 (in comparison with proCOL5A1 and proCOL11A2). In addition, the expression of the proCOL11A1 gene was measured by RT-qPCR. On the other hand, the expression of proCOL11A1 was compared to the expression of estrogenic receptors, progestagen receptors, the state of the epidermal growth factor receptor 2 (HER2), the histologic grade and the stage of the disease. We also compared the immunohistochemical expression of proCol11A1 to the disease-free interval, and to overall survival. Results: The immunohistochemical analysis showed that proCOL11A1 was expressed in 100% of infiltrating ductal carcinomas, but only focally expressed in 2.2% (1 case) of sclerosing adenosis, in agreement with RT-qPCR results. ProCOL11A1 expression did not prove to have a prognostic value in relation to the disease-free interval or to overall survival in infiltrating ductal carcinoma. Conclusion: The anti-proCOL11A1 pAb is a stromal marker for breast cancer and the expression of proCOL11A1 does not seem to have a prognostic value in infiltrating ductal carcinoma of the breast.
- PublicationOpen AccessElucidation of soft tissue flap histologic margins within a canine vocal fold(F. Hernández y Juan F. Madrid. Universidad de Murcia: Departamento de Biología Celular e Histología, 2015) Glab, Rachel C.; Gunderson, McLean; Torrealba, José; Bauer, Ben; Dailey, Seth H.Summary. Background: Histologic identification of implanted soft tissues in experimental animal models can be challenging, as donor tissue often strongly resembles the recipient bed. We have encountered this dilemma following implantation of a Composite Thyroid Ala Perichondrium flap (CTAP) into a vocal fold. The CTAP procedure is the first to utilize a vascularized flap for vocal fold reconstruction, making data to confirm or refute its viability critical. The current study evaluated several tissue stains to define precisely the histologic margins of CTAPs at two weeks post-implantation in a canine model. Methods: Initial testing exposed canine cadaveric tissues to four stains (tattoo ink, Congo red, 4’6- diamidino-2-phenylindole, and henna) across four time periods. Tattoo ink alone withstood histologic processing. An exposure of 1 minute adequately delineated CTAP boundaries. The study concluded with a canine in vivo evaluation of a CTAP exposed to tattoo ink for 1 minute. After a two-week recovery period, vocal folds were harvested and evaluated histologically. Results: Tattoo ink proved to be a safe and effective histologic marker in vivo, where the histologic margins of the implanted CTAP were clearly demarcated by a thin band of tattoo ink, soft tissue reactions were minimal, and interference with standard, special, or immunohistochemical stain assessments did not occur. Conclusions: Tattoo ink provides a reliable means of demarcating a CTAP within a vocal fold and demonstrated that CTAPs survive transplantation. Further, tattoo ink demarcation may serve as a useful histologic marker for those wishing to assess tissue implants in other in vivo models.
- PublicationOpen AccessExpression of matrix Gla protein and osteocalcin in the developing tibial epiphysis of mice(F. Hernández y Juan F. Madrid. Universidad de Murcia: Departamento de Biología Celular e Histología, 2015) Liu, Hongrui; Guo, Jie; Wei, Shanliang; Lv, Shengyu; Feng, Wei; Cui, Jian; Hasegawa, Tomoka; Hongo, Hiromi; Yang, Yang; Li, Xiangzhi; Oda, Kimimitsu; Amizuka, Norio; Li, MinqiThis study aimed to investigate the expression of matrix Gla protein (MGP) and osteocalcin (OCN) in the tibial epiphysis of developing mice. At 1, 2, 3, and 4 weeks after birth, tibiae were removed and processed for histochemical observations and western blot analyses under anesthesia. To evaluate bone volume, the specimens were scanned with Micro CT Scanner from the articular cartilage through the growth plate, along the long axis of tibia. At 1 week after birth, OCN reactivity was faint in the region of vascular invasion, while hardly any MGP reactivity was discernible. Subsequently, MGP reactivity was seen on the cartilaginous lacunar walls of hypertrophic chondrocytes, while OCN reactivity was evenly found not only in the bone matrix, but also in the cartilaginous lacunar walls and on the bone surfaces. Furthermore, double-immunostaining clearly showed that MGP reactivity appeared closer to the cartilage matrix than OCN reactivity until postnatal week 3. Interestingly, the immunoreactivities for MGP and OCN both showed tidemarks in the articular cartilage at postnatal week 4, and MGP reactivity was more intense than OCN reactivity. Statistical analyses showed an overall upward trend in MGP and OCN expression levels during tibial epiphysis development, even though OCN was more abundant than MGP at every time-point. Taken together, our findings suggest that the expression of MGP and OCN increased gradually in the murine developing tibial epiphysis, and the two mineral-associated proteins may occur at the same location during a particular period, but at different levels.
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