Alpha basic crystallin expression in canine mammary tumors

The aim of this study was to evaluate prognostic and/or diagnostic factors of canine mammary tumors by immunohistochemically analyzing the expression of alpha basic crystallin (alphaB-c). For this, formalin-fixed, paraffin-embedded blocks of 51 naturally-occurring canine mammary tumors (11 benign and 40 malignant) were used. Tissue from eight normal canine mammary glands were served as a control. Immunohistochemically, in the control mammary tissues, a few luminal epithelial cells were alphaB-c positive but myoepithelial cells were negative. In benign or simple type malignant tumors, alphaB-c expression was observed in luminal epithelial cells while the myoepithelial basal cells were negative. In benign or complex type malign tumors, positive staining was predominantly found in the cytoplasm of epithelial cells. Immunoreactivity of alphaB-c was also observed in neoplastic myoepithelial cells. Statistically, the number of cells immunolabeled with alphaB-c was found to be significantly different among tissues from normal canine mammary glands, benign lesions, and malignant tumors (p < 0.05). alphaB-c immunoreactivity was higher in malignant tumors than the control mammary tissues (p < 0.001). Data obtained in the current study revealed a strong association between high expression levels of alphaB-c and primary mammary gland tumors in canines.

alphaB-Crystallin is a Novel Oncoprotein Associated with Poor Prognosis in Breast Cancer / 한국유방암학회지

PURPOSE: alphaB-crystallin, a small heat shock protein, is an anti-apoptotic protein associated with aggressive tumor behavior. A recent study revealed that alphaB-crystallin is overexpressed in a metastatic variant of the GI101A human breast carcinoma cell line. The purpose of this study was to investigate whether alphaB-crystallin is related to other breast tumor markers and can predict a breast cancer prognosis. METHODS: Eighty-two patients who underwent breast cancer surgery at Hallym Sacred Heart Hospital were enrolled. alphaB-crystallin expression was determined by immunohistochemical staining. Estrogen receptor, progesterone receptor (PR), human epidermal growth factor receptor, lymphovascular invasion, histological grade, other tumor markers and time to recurrence were compared with alphaB-crystallin expression. RESULTS: alphaB-crystallin expression in breast cancer tissues was associated with PR (p=0.030), the number of metastatic lymph nodes (pN) (p=0.020), lymphovascular invasion (p=0.022), histological grade (p=0.004) and triple negative breast cancer (TNBC) (p=0.004). alphaB-crystallin expression significantly decreased time to recurrence (p=0.039). CONCLUSION: The results revealed a strong relationship between alphaB-crystallin and poor prognostic factors such as the number of metastatic lymph nodes (especially pN2), TNBC, and rapid time to recurrence. We believe that alphaB-crystallin could be a novel oncoprotein biomarker of a poor prognosis in breast cancer.

Characterization of the antigenic phenotype of alphaB-crystallin-expressing peripapillary glial cells in the developing chick retina / 대한해부학회지

Radial glia are transdifferentiated into astrocytes within the developing brain and spinal cord. The neural retina contains Muller cells, which are retinal radial glia. Some of the cells that surround the optic nerve head among Muller cells in the chicken retina are called peripapillary glial cells (PPGCs). PPGCs express different molecules compared to typical Muller cells. However, an antigenic PPGC phenotype has not yet been clearly established. In this study, we classified the antigenic PPGC phenotypes and identified the differentiation stages of these cells. At embryonic day (E)8, alphaB-crystallin-positive PPGCs had a bipolar shape with long processes that traversed entire layers of the retina. Pax2 and vimentin were expressed in alphaB-crystallin-positive PPGCs. Glial fibrillary acidic protein (GFAP) immunoreactivity was not observed in PPGCs. At E18, alphaB-crystallin immunoreactivity disappeared from the vitread processes of PPGCs. However, the PPGC cell bodies and ventricular processes contained alphaB-crystallin protein, and the PPGCs retained the same Pax2-positive/vimentin-positive/GFAP-negative profile as that seen at E8. At post-hatch day 120, alphaB-crystallin and Pax2 immunoreactivity was not observed, but vimentin and GFAP expression was clearly observed in the presumptive location of the PPGCs. Furthermore, these two proteins overlapped within that location. Considering that vimentin expression is prolonged until the post-hatching period in chicken brain, these findings suggest that Pax2-negative/vimentin-positive/GFAP-positive PPGCs are phenotypically identical to mature astrocytes in this avian species.

Detection of alphaB-crystallin mRNA using Single-stranded DNA Probe in Oligodendrocytes of the Developing Chick Retina / 체질인류학회지

In situ hybridization (ISH) using single-stranded DNA probe (ssDNA probe) is a useful method for observing the specific transcripts in cells, since it is convenient to prepare probe which is specific and sensitive. In this study, ssDNA probe for detection of alphaB-crystallin (aBC) mRNA, transcript of a heat shock protein, was prepared and aBC mRNA-expressed cells were spatiotemporally observed in the retina of the developing chick embryos. Single-stranded antisense probe produced by reverse transcription and polymerase chain reaction was identified as a specific probe for aBC mRNA in comparison to negative control using sense probe and immunohistochemistry for aBC protein. In the ISH experiment, aBC mRNA was expressed only in the peripapillary glial cells which are a specific cell type located in the avian retina adjacent to the optic nerve at E12 and E14 retinas. At E16, a small number of aBC mRNA-expressed cells were identified in the nerve fiber layer (NFL) of the retina. At E18, aBC mRNA-expressed cells were observed in the ganglion cell layer (GCL) as well as the NFL. At E20, the number of aBC mRNA-expressed cells was increased in the GCL and the NFL. Based on the same localization of nkx2.2 immunoreactive cells and aBC mRNA-expressed cells, aBC mRNA-expressed cells were identified as oligodendrocytes. These results indicate that ssDNA probe for aBC mRNA detection is very useful tool for oligodendrocyte research such as distribution, migration and differentiation of the cells.

Gene Knockdown in the Olfactory Bulb, Amygdala, and Hypothalamus by Intranasal siRNA Administration / 대한해부학회지

Intranasal administration provides a method of bypassing the blood brain barrier, which separates the systemic circulating system and central interstitial fluid, and directly delivering drugs to the central nervous system. This method also circumvents first-pass elimination by the liver and gastrointestinal tract. In the present study, the authors investigated intranasal siRNA delivery efficiency by using FITC-labeled transfection control siRNA and a genespecific siRNA. The localization of fluorescence-tagged siRNA revealed that siRNA was delivered to cells in the olfactory bulb and that the level of the siRNA target gene (alpha B-crystallin) was significantly reduced in the same area. siRNA was delivered to processes as well as nuclei and cytoplasm. At 12 hrs after intranasal delivery, siRNA-mediated target gene reduction was observed in other more distally located brain regions, for example, in the amygdala, entorhinal cortex, and hypothalamus. Target gene knockdown was demonstrated by double immunohistochemistry, which demonstrated alpha B crystallin expression depletion in more than 70% of cells at 12 hrs after the intranasal delivery. siRNA-mediated target gene suppression was detected not only in neurons but in glia, for example, astrocytes. These results indicate that intranasal siRNA delivery offers an efficient means of reducing specific target genes in certain regions of the brain and of performing gene knockdown-mediated therapy.

Expression of HSP27 and alphaB-crystallin in Avian Cerebellum during Development / 대한해부학회지

It is well known that small heat shock proteins play a role as molecular chaperone. However, during normal development of the cerebellum, expression and distribution of HSP27 and alphaB-crystallin (alphaBC) which are small heat shock proteins have not been reported. To verify the protective role of HSP27 and alphaBC in neurons and glial cells, we examined the expression and distribution of HSP27 and alphaBC in the developing chick cerebellum using immunoblot, immunohistochemical and double immunofluorescence staining. Expression of both HSP27 and alphaBC was first identified in the cerebellum of the embryonic day 14 (E14) embryo, and was increased at E18. Double immunofluorescence analysis with myelin-basic protein (MBP) demonstrated that alphaBC positive (+) cells were mature myelinating oligodendrocytes. alphaBC+ cells were observed in the white matter of the E14 cerebellum. At E18, there were a number of alphaBC+ cells in the white matter and a few cells in the granular layer of the gray matter. On the other hand, HSP27+ cells were observed in the white matter and the Purkinje cell layer at E14. At E18, HSP27+ signals were observed in Purkinje cells and neurons of cerebellar nucleus as well as oligodendrocytes in the white matter and the granular layer. The results that HSP27 and alphaBC were expressed in specific neurons and glial cells in the developing cerebellum suggest that HSP27 and alphaBC may be involved in the protective mechanism for the apoptosis of neurons and the physiological stress occurred in oligodendrocyts during cell maturation.

Induction of alphaB-crystallin in the hippocampus of KA-treated mouse brain / 대한해부학회지

The alphaB-crystallin, which is a member of small heat shock protein (sHSP), was initially identified as a component of a vertebrate eye lens protein, and also shown to be expressed in non-lenticular tissues including cardiac muscles and the central nervous system. Recently, it was demonstrated that alphaB-crystallin expression was increased in the brain of patients suffering various neurological diseases including Alzheimers disease. In the current study, we examined in detail the time-course of alphaB-crystallin expression in the region of neuronal loss and in activated glia cells in hippocampus of the KA-treated mouse brain. The alphaB-crystallin was expressed in pyramidal layer and in oligodendrocytes of hippocampus 1 day after KA-treatment, which was similar to that in normal mice. The alphaB-crystallin expression began to be increased 2 days after KA-treatment and reached peak induction, especially in astrocytes in the CA3 area of hippocampus 4 days after KA-treatment. Immunofluorescent staining with anti-alphaB-crystallin and anti-GFAP antibodies revealed that the induction of alphaB-crystallin was localized in the activated astrocytes, prominently in the CA3 region of hippocampus where a severe neuronal death was undergoing. The results suggested that alphaB-crystallin might play a role in reactive gliosis and/or in delayed neuronal death proceeded in KA-induced epileptic brain.