Histology and histopathology Vol.32, nº6 (2017)

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Open Access
Immunostaining of proinflammatory cytokines in renal cortex and medulla of rats exposed to gold nanoparticles
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Khan, Haseeb A.; Ibrahim, Khalid E.; Khan, Ayaat; Alrokayan, Salman H.; Alhomida, Abdullah S.
Recently, gold nanoparticles (GNPs) have shown promising applications in targeted drug delivery and contrast imaging. Although in vitro cytotoxicity of GNPs has been thoroughly studied, there are limited data on in vivo toxicity of GNPs. In this study, we evaluated the effects of intraperitoneally injected 10 nm and 50 nm GNPs (5 µg/animal) on the expression of proinflammatory cytokines (IL-1β, IL-6 and TNF-α) on day 1 and day 5, post-exposure. The results of immunohistochemistry showed that both 10 nm and 50 nm GNPs induced an acute phase expression of proinflammatory cytokines in renal cortex and medulla. This proinflammatory response was comparatively more intense in renal medulla than cortex. All the three cytokines were undetectable in control cortex and medulla. In conclusion, both 10 nm and 50 nm GNPs caused an acute phase induction of proinflammatory cytokines in cortex and medulla of rat kidneys. An intense immunostaining of proinflammatory cytokines in renal medulla warrants further studies to evaluate the nephrotoxicity of GNPs to validate the safe application of GNPs for contrast imaging in renal insufficiency.
Open Access
Dynamin-related protein 1 (Drp1) mediating mitophagy contributes to the pathophysiology of nervous system diseases and brain injury
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Wu, Qiong; Luo, Cheng Liang; Tao, Lu Yang
As the main source of energy (celluar ATP) in eukaryotic cells, mitochondria are involved in cellular physiology and pathology. The balance of mitochondrial dynamic, fission and fusion regulated by quality control mechanisms, provides a guarantee for maintaining mitochondrial function, even celluar function. Worn out mitochondria would be removed through mitophagy which is regulated by autophagy related proteins and mitochondrial membrane proteins. Drp1, dynamicrelated protein 1, is regarded as one of the most important proteins to evaluate mitochondrial fission mediating mitophagy in neurodegenerative diseases (eg. Alzheimer’s, Parkinson’s, Huntington’s, amyotrophic lateral sclerosis) and heart failure. Recent studies have focused on the roles of Drp1 in ischemia-induced mitophagy in the hippocampal CA3 region, and traumatic brain injury (TBI)-induced cell death together with functional deficits. However, the exact mechanisms have not been well characterized. In this review, we will discuss and clarify the role of Drp1 and mitophagy in nervous system diseases and brain injury therein, with a special emphasis on their molecular mechanisms mediating mitochondrial dynamics and mitophagy
Open Access
Immunohistochemical expression of mucin antigens in gallbladder adenocarcinoma: MUC1-positive and MUC2-negative expression is associated with vessel invasion and shortened survival
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Hiraki, Tsubasa; Yamada, Sohsuke; Higashi, Michiyo; Hatanaka, Kazuhito; Yokoyama, Seiya; Kitazono, Ikumi; Goto, Yuko; Kirishima, Mari; Batra, Surinder K.; Yonezawa, Suguru; Tanimoto, Akihide
Mucins play pivotal roles in influencing cancer biology, for example affecting carcinoma invasion, aggressiveness and/or metastatic potential. Our aim is to investigate the significance of expression profiles of two mucins in particular, MUC1 and MUC2, their correlations with various clinicopathological features, and prognosis in gallbladder adenocarcinoma (GBAC). We performed immunohistochemistry from patients with surgically resected GBAC, using antibodies against mucin core proteins MUC1/DF3 and MUC2/Ccp58 in 81 paraffin-embedded tumor samples. MUC1 or MUC2 expression was considered to be high when ≥20% or 10% of the GBAC cells showed positive staining, respectively. High MUC1 expression was revealed to have a significant relationship to the presence of pathologically lymphatic and vascular invasion, and regional lymph node metastasis. By contrast, high MUC2 expression showed a significant correlation with pathologically perineural invasion, T stage ≥3, and post-operative recurrence. Moreover, MUC1 showed significantly positive co-expression and potentially complementary correlations with MUC2. Multivariate analyses demonstrated that the high MUC1 expression group had significantly shorter diseasespecific survival times. However, the combination of both high MUC1 and MUC2 expression did not predict worse outcome in GBACs. Therefore, although each mucin has a somewhat important role in the pathogenesis of GBAC progression, MUC1 can independently predict vessel invasion and poor prognosis in patients with GBAC. The detection of MUC1 might well offer a useful parameter for providing clinical management and treatment against postsurgical GBACs.
Open Access
The effect of bilberries on diabetes-related alterations of interstitial cells of Cajal in the lower oesophageal sphincter in rats
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Velickov, Aleksandra; Mitrovic, Olivera; Djordjevic, Branka; Sokolovic, Dusan; Zivkovic, Vladimir; Velickov, Asen; Pantovic, Vukica; Poklar Urlih, Natasa; Radenkovic, Goran
Diabetic gastroenteropathy involves not only the parasympathetic and sympathetic autonomic nerves, but also enteric neurons, smooth muscle cells and interstitial cells of Cajal (ICC). ICC are the cells of mesenchymal origin that occur within and around the muscle layers in the gastrointestinal tract. The objective of the present study was to investigate the alterations of ICC in the lower oesophageal sphincter (LOS) of streptozotocin-nicotinamide non-insulin-dependent diabetes rats. Moreover, we investigated possible ICC in rats with the same type of diabetes, treated with bilberry fruit extract, bearing in mind that its hypoglycemic effect had been already proven. Male Wistar rats (10 weeks old) were used, and diabetes was induced by an intraperitoneal injection of streptozotocin, immediately after intraperitoneal application of nicotinamide. The specimens were exposed to anti-c-kit antibodies to investigate the distribution of ICC, and the smooth muscle cells were immunohistochemically labelled using anti-desmin antibodies. Intramuscular ICC were very abundant in the LOS of rats. They were spindle-shaped, with two long processes connecting them into long linear sequences. In the LOS of diabetic rats, intramuscular ICC were rarely present and linear cell-cell connections between these cells were completely missing. In groups treated with bilberry, the number and distribution of ICC were exactly the same as in the above described rats with induced diabetes. In summary, a decrease of intramuscular ICC, discontinuities and breakdown of contacts between ICC were observed in streptozotocin-nicotinamide induced diabetes rats and in groups treated with bilberry. Bilberry fruit extract was shown to have hypoglycemic activity, but without any protective effects on ICC in the LOS of diabetic rats.
Open Access
Glucose-mediated cytoprotection in the gut epithelium under ischemic and hypoxic stress
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Huang, Chung Yen; Pai, Yu Chen; Yu, Linda Chia Hui
Single-layered intestinal epithelia play key roles in the maintenance of gut homeostasis and barrier integrity. Various types of epithelial cell death, including apoptosis, necrosis, and necroptosis, have been detected in ischemic and hypoxic stress conditions, thus resulting in bacterial translocation and gut-derived septic complications. Cytoprotective strategies, such as enteral glucose uptake, rescue intestinal epithelium from cell death after ischemic and hypoxic injury. Although glucose metabolism and energy production are generally considered to be the key factors in cytoprotection, the precise modes and sites of action have not been clarified. Our recent studies have demonstrated that energy restoration promotes crypt hyperplasia but does not prevent epithelial cell death under ischemic stress. On the other hand, glycolytic pyruvate prevents epithelial cells from undergoing apoptosis and necroptosis by scavenging free radicals in an ATP-independent manner. Distinct gut protective mechanisms involving ATP, pyruvate, glucose metabolic enzymes, and sodiumdependent glucose transporter activation are discussed here. Overall, glucose-mediated cytoprotection may be a universal mechanism that has evolved in epithelial cells for the maintenance of intestinal homeostasis. Enteral glucose supplementation is beneficial as a perioperative supportive therapy for the protection of gut barrier integrity.
Open Access
Significant up-regulation of 1-ACBP, B-ACBP and PBR genes in immune cells within the oesophageal malignant tissue and a possible link in carcinogenic angiogenesis
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Dlamini, Zodwa; Mbele, Mzwandile; McCabe, Michelle; Rees, Jasper; Naicker, Saraladevi; Mbita, Zukile
Oesophageal cancer ranks as the sixth most common malignancy in the world, and recent evidence has shown that its incidence is increasing. ACBPs (AcylcoA binding proteins) act as intracellular carrier-proteins for medium to long chain acyl-coA, mediating fatty acid transport to the mitochondrion for β-oxidation. ACBPs are also believed to be putative ligands of PBR (peripheral benzodiazepine receptor), and once they bind to this receptor they facilitate mitochondrial membrane permeabilization, presumably favouring apoptosis. The main aim of the study was to establish the expression patterns of 1- Acyl-coA binding proteins (1-ACBP), BAcyl-coA binding proteins (B-ACBP), and peripheral bezodiazepine receptor (PBR) in oesophageal cancer, and to link their roles with the disease. In situ hybridization and quantitative real-time PCR methods were performed to determine localization and the expression levels of the three genes in oesophageal cancer. All three genes products illustrated substantial up-regulation within the malignant tissue sections as compared to normal oesophageal sections. All three transcripts localized specifically to mast cells, plasma cells and lymphocytes in diseased and normal tissue section. In the diseased tissue, B-ACBP and 1-ACBP mRNA localized to endothelial cells of blood vessels in the submucosa. B-ACBP also localized to the nucleus of squamous epithelial cells. PBR localization was found in tumour islands of invasive tissue sections. Quantitative RT-PCR also indicated that the expression levels of PBR were higher as compared to the ACBP genes expression in tumours. These results show that 1-ACBP, B-ACBP and PBR play a role in the pathogenesis of oesophageal tumours and possibly in carcinogenic angiogenesis.
Open Access
Morphometric characterization of the human portal and hepatic venous trees: A quantitative support to the liver micro-anatomic models free of subunits
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Almenar Medina, Sergio; Palomar De Lucas, Brenda; Guerrero Albors, Ester; Ruiz Sauri, Amparo
Conventional models of liver microanatomy assume the presence of subunits. Nevertheless, some researchers propose that the liver is a continuous structure, free of these subunits, but with a characteristic vascular pattern. The present study describes a morphometric analysis of portal and hepatic veins in 50 human autopsy non-pathological liver samples. The main objective was to measure three proportions: 1. portal tracts / hepatic veins, 2. distributing portal veins / distributing hepatic veins and 3. terminal portal veins / terminal hepatic veins. These ratios were compared with the traditional microcirculatory liver models. Our material comprised 3,665 portal veins and 3,761 hepatic veins. The minimum diameter of half of the venous vessels of both types belongs to the interval (25 μm , 60 μm), given that 1881 portal veins (49.434%) and 1924 hepatic veins (50.565%) fall within this interval. We have statistically shown with the χ2 test (α=0.990) that the portal and hepatic veins belonging to the interval (25 μm, 400 μm) (distributing veins) had an identical proportion. If the portal and hepatic veins are arranged according to the principle of interdigitation of Takashasi (1970), there should be an almost identical number of both types of veins. Our results contradict the presumably numeric preponderance of distributing portal veins with regard to the distributing hepatic veins that is inherent in the models of Kiernan, Matsumoto and Rappaport.
Open Access
Localization of choline acetyltransferase and tyrosine hydroxylase immunoreactivities in the superior colliculus of the microbat, Rhinolophus ferrumequinum
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Jeong, Se Jin; Jeon, Chang Jin
The purpose of this study was to determine whether the superior colliculus (SC) of the microbat has the same neurochemical makeup as that of other mammals. We examined the organization of choline acetyltransferase (ChAT)- and tyrosine hydroxylaseimmunoreactive (TH-IR) fibers/cells using standard immunohistochemistry with antibodies against ChAT and TH. ChAT-IR fibers observed in the superficial layers were denser than those in the deeper layers, and these fibers were classified into two types: small varicose fibers and large varicose fibers. ChAT-IR cells were predominantly located in the superficial layers with diverse morphologies. Among the well-known sources of cholinergic fibers in the mammalian SC, pedunculopontine tegmental nucleus (PPTN) and laterodorsal tegmental nucleus (LDTN) contained strongly labeled ChAT-IR cells, while no cholinergic structures were found in the parabigeminal nucleus (PBG) in the microbat brain. TH-immunoreactivity was found within fibers but not within cells. The density of TH-IR fibers was high in the zonal layer, moderate in the superficial gray and optic layers, and low in the deeper layers. Well-labeled TH-IR cells were also observed within area 13 and the locus coeruleus, known as the sources of catecholaminergic fibers in other mammalian SC. Although there are some cytoarchitectural variations among species, our results clearly showed elaborately organized ChAT-IR and TH-IR fibers/cells in the microbat SC. Our findings will contribute significantly to the understanding of actively constructed microbat visual systems
Open Access
CORRIGENDUM TO: "High glucose concentration-induced expression of pentraxin-3 in a rat model of continuous peritoneal dialysis" Histol Histopathol. 2016 Nov;31(11):1251-1258. doi: 10.14670/HH-11-756]
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Ishimatsu, Nana; Miyamoto, Tetsu; Ueno, Hiromichi; Hasegawa, Emi; Kuma, Akihiro; Fujimoto, Yoko; Bando, Kenichiro; Nakamata, Junichi; Furuno, Yumi; Serino, Ryota; Baba, Ryoko; Morimoto, Hiroyuki; Doi, Yoshiaki; Tamura, Masahito; Otsuji, Yutaka
Open Access
Inflammatory mediators and signalling pathways controlling intervertebral disc degeneration
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Navone, Stefania Elena; Marfia, Giovanni; Giannoni, Amedeo; Beretta, Matteo; Guarnaccia, Laura; Gualtierotti, Roberta; Nicoli, Daniele; Rampini, Paolo; Campanella, Rolando
Intervertebral disc (IVD) degeneration (IDD) is one of the major causes of back pain, a condition that represents a serious socio-economic burden. Deeper knowledge of the complex and fine relationship between IVD degeneration, tissue inflammation and pain, appears to be critical to improve the current therapies, which have so far proven themselves ineffective. Upon degeneration, IVD tissues become inflamed, and this inflammatory microenvironment is associated with a cascade of degenerative events that may eventually cause discogenic pain. In particular, several studies have highlighted the major role of a number of proinflammatory mediators not only in the onset of the inflammatory condition, but also in the development of IDD in general. In this review, we will present the main pathological events that occur during disc degeneration, focusing on the relationship between the abnormal inflammatory milieu of the degenerating IVD, IDD and the generation of pain. Finally, we will present the current therapies for the treatment of IDD and low back pain, and the perspectives of future, more effective therapies.
Open Access
Ghrelin and adipose-derived mesenchymal stromal cells improve nerve regeneration in a rat model of epsilon-caprolactone conduit reconstruction
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Hernández Cortés, Pedro; Toledo Romero, Miguel Angel; Delgado, Mario; Gonzalez Rey, Elena; Gómez Sánchez, Rafael; Prados Olleta, Nicolás; Aneiros Fernández, José; Crespo Lora, Vicente; Aguilar, Mariano; Galindo Moreno, Pablo; O’Valle, Francisco
Objective. Attempts have been made to improve nerve conduits in peripheral nerve reconstruction. We investigated the potential therapeutic effect of adipose-derived mesenchymal cells (ASCs) and ghrelin (GHR), a neuropeptide with neuroprotective, trophic, and developmental regulatory actions, on peripheral nerve regeneration in a model of severe nerve injury repaired with nerve conduits. Material and methods. The right sciatic nerves of 24 male Wistar rats were 10-mm transected unilaterally and repaired with Dl-lactic-ε-caprolactone conduits. Rats were then treated locally with saline, ASCs, or GHR. At 12 weeks post-surgery, we assessed limb function by measuring ankle stance angle and percentage muscle mass reduction and evaluated the histopathology, immunohistochemistry, ultrastructure, and morphometry of myelinated fibers. Main Results. Rats receiving GHR or ASCs showed no significant increased functional recovery in ankle stance angle (p=0.372) but a higher nerve area (p=0.015), myelin area (p=0.046) and number of myelinated fibers (p=0.012) in the middle and distal segments of operated sciatic nerves in comparison to saline-treated control animals. Conclusion. These results suggest that utilization of ghrelin or ASCs may improve nerve regeneration using Dl-lactic-ε-caprolactone conduits.

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