Determining Role of TGF-β3 in Some Treatments of Albino Mouse, Mus musculus Linnaeus, 1758 Infected with Hydatid Cyst by Using an Immunohistoflourescent Staining Technique

The present study was conducted as an attempt to treat secondary hydatid disease in white mice strain Balb/c by immunohistoflourescent (IHF) staining technique. For this purpose, the mice were infected with hydatid cysts and treatment was done by oxfendazole (OFZ) at a concentration of 30 mg/kg, praziquantel (PZQ) at a concentration of 40 mg/kg and albendazole (ABZ) at a concentration of 10 mg/kg of body weight. Each drug was given weekly for four months and the same concentrations as above. The results showed that the highest treatment efficiency was in case of OFZ + PZQ treated mice, while the groups treated with OFZ, OFZ + ABZ and ABZ + PZQ showed less treatment efficiency, respectively. The IHF staining technique was used to determine the cytokinesis of TGF-β3 in the spleen and liver of experimental mice. For this reason, OFZ is considered as one of the most promising chemotherapies used in the treatment of hydatid cysts.

Macro and Micro Anatomical Observation on the Testes of Local Dog (Canis lupusfamiliaris) of Assam

Background: The study on testes of local dog of Assam is of great value in regard to germplasm conservation. The aim of the study was to evaluate the gross and histomorphological examination of testes of male reproductive system. Methods: The testes were collected at the time of castration from Department of Surgery and Radiology, College of Veterinary Science, Assam Agricultural University, Khanapra, Guwahati, Assam, India. The research was carried out for a period of one year in Department of Anatomy and Histology, College of Veterinary Science, Assam Agricultural University, Khanapara, Guwahati, Assam. Then gross anatomical studies were made on it and the tissue samples were fixed in 10% neutral buffered formalin solution and were processed as per the standard technique of procedure (Luna, 1968). The paraffin blocks were sectioned in Shandon Finesse microtome at 5 µm thickness and the sections were stained with Mayer’s Haematoxylin and Eosin staining technique for Cellular details as per the method of Luna (1968). Result: Grossly, the testes of local dog consisted of two surface viz., lateral and medial and two ends i.e. upper end and lower end. The upper end of the testes was occupied by the head of the epididymis and the lower end of the testes was occupied by the tail of the epididymis. Mediastinum testis was observed in the centre of testes of local dog. Histologically, the testes were covered by serous layer (Tunica vaginalis), connective tissue layer (Tunica albugenia) and vascular layer (Tunica vasculosa) from outside to inwards. Spermatogenic cells like spermatogonia, primary spermatocytes, secondary spermatocytes, spermatids and spermatozoa, and sertoli cells were observed in seminiferous tubules. The sertoli cells were attached to the basement membrane of seminiferous tubules. Cluster of Leyding cells were found between the semineniferous tubules and it contained large spherical nuclei. The epididymides were lined by pseudo stratified ciliated columnar epithelium.

Evaluation of an improved Giemsa staining technique for blood borne parasite in thick and thin blood film from dogs

Many a times the use of rapid diagnostic tests for blood borne parasites like trypanosomiasis and Babesiosis is increasingly being used, but the gold standard for its detection is still the use of microscopy both as reference and confirmatory diagnosis. To determine the effectiveness of the adjusted stock giemsa staining technique over the conventional methods. Venous Blood samples were collected from 10 dogs in EDTA and then used for the simultaneous preparation of two thin and thick smear slides, one stained according to Giemsa 1:20 dilution for 30 mins while the other was stained using the Stock Solution for 30seconds the diluted with buffered saline for 20seconds and rinsed. Fixation of the thin smear was done in a covered staining jar containing anhydrous methanol for 1 to 2 min, after which the slides were air-dried. From the result obtained from 10 dogs blood samples gotten from the veterinary clinic, the adjusted giemsa staining technique showed a positive differentiation when compared to the 1:20 dilution, a total of 7 blood samples tested positive for blood borne parasites, Trypanosoma evansi, Babesiosis cani and Heart worm. The highest percentage occurrence was T.evansi (40%), Babesiosis cani(20%) and Heart worm (10%).The adjusted Giemsa staining technique serves as a fast, easy and less complex alternative to the 1:20 dilution, where the solution has to be diluted from the stock solution and then stained, although the only disadvantage to this technique would be easy contamination of the stock solution, but the advantages here is that it saves time, quicker result output and better differentiation microscopically.

Anticancer and Gene Expression Analysis of Piper nigrum Extract on Colon Cancer Cell Lines

One of the most prevalent malignancies among geriatrics is colorectal cancer (CRC), which starts to develop in the forms of genetic syndromes in young adults. The Piper nigrum is one important common spice used in the household having anticancer activities. The current study aims to evaluate P. nigrum seed extracts potency as anticancer against CRC cell line (COLO205). The extract is used to elucidate the MTT assay, DNA damage studies (COMET assay), Acridine Orange/Ethidium Bromide dual staining, cell death, cell cycle arrest using Flow cytometry, and regulation of Bcl-2, Bax & P53 gene regulation. To check the cell cytotoxicity by MTT assay methanolic extract was used. To evaluate anticancer activity the sample was extracted in methanol. RT-PCR was used to elevate gene expression studies of Bcl-2, Bax, and P53. In the dose-dependent mode, the extract inhibited the growth of COLO205 cells and the IC50 value was calculated at 48.2 μg/ml. The DNA fragmentation induced by apoptosis was the primary reason for the cell toxicity as observed by DNA damage studies & AO/EB dual staining technique. The extract concentration ranging from 40 & 80 μg/ml remarkably increased the proportion of cells in the S & G2/M phase. Cells at the late-apoptotic stage were found to be in the range of 22% - 57%. The Bax and P53 were upregulated and Bcl-2 was downregulated when treated with the extract. From this investigation underlying the mechanism of CRC was found to be P. nigrum extract caused to induce apoptosis and upregulation of tumor suppressor gene downregulation of apoptosis-suppressing gene bcl-2.

Gross, Histomorphological and Scanning Electronic Microscopic Studies on Gut Associated Lymphoid Tissue of Intestine of Pati Duck (Anas platyrhynchos domesticus) of Assam

Background: The study on Gut Associated Lymphoid Tissue (GALT) of Pati duck of Assam is of great value in regard to normal academic and bio-medical research aspects. The aim of the study was to evaluate the gross, histomorphological and scanning electronic microscopic examination of gut-associated lymphoid tissue of the intestine of Pati duck at different age group. Methods: For this study, forty five Pati ducks were divided into five groups depending on its age viz., 1st week, 4th week, 16th week, 24th week and 42nd weeks old. The pieces of gut having lymphoid tissue or Peyer’s patches were collected immediately after slaughter. These samples were fixed in 10% neutral buffered formalin solution and were processed as per the standard technique of procedure (Luna, 1968). The paraffin blocks were sectioned in Shandon Finesse microtome at 5 µm thickness and the sections were stained with Mayer’s Haematoxylin and Eosin staining technique for Cellular details, Van Gieson’s method for collagen fibres, Gomori’s method for reticular fibres, Hart’s method for elastic fibres and Bielchowsky’s method for axis cylinder and dendrites as per the method of Luna (1968). Result: Gut-associated lymphoid tissue was found in the duodenum, jejunum, ileum, caecum and the terminal part of the rectum in all the age group of Pati duck. The lymphoid compartment of the gut-associated lymphoid tissue in duck included a follicular structure, dome, follicle associated epithelia and interfollicular area. Lamina propria of jejunum was heavily infiltrated with diffuse lymphatic tissue in the 16th, 24th and 42nd week of age of Pati duck. The scattered and diffuse lymphatic infiltration occurred in all age groups. The lamina propria of the colorectum revealed an extensive network of reticular fibre with diffused lymphatic tissue in all the age group of duck. In Scanning Electron Microscope, the lumen of the jejunum was covered by finger-like villi with numerous opening of goblet cells. The lymphoid follicle of Lamina propria contains numerous lymphocytes along with connective tissue fibres.

Investigating the Mechanism Behind the Release of Microcystins in Freshwater Cyanobacteria

<p>The frequency and distribution of toxic cyanobacterial blooms are increasing globally, creating the need for a better understanding of the processes involved in toxic secondary metabolite production. Microcystins (MCs) are potent hepatotoxins produced by a wide range of bloom-forming cyanobacteria genera such as Microcystis and Planktothrix. Although the release of MCs to the extracellular environment has long been considered a by-product of cell lysis and death, several studies suggest the presence of a mechanism that actively transports these toxins outside the cell membrane. The aim of the present study was to find evidence for a link between cell lysis and concentrations of extracellular MCs. A dual-fluorescence cell viability assay using the nucleic acid stain SYTOX Green was optimised for use on Microcystis and Planktothrix. A SYTOX Green concentration of 1 µM, and an incubation time of 30 minutes, yielded a bright and even fluorescent signal that readily identified lysed cells. The improved staining technique, in conjunction with liquid chromatography-mass spectrometry analyses, was employed in a culturing experiment to track the transfer of MCs to the extracellular environment in relation to the amount of cell lysis. For Microcystis, there was a strong and significant positive relationship between cell lysis and the concentration of extracellular MC. When the extracellular MC was predicted according to cell lysis levels and the MC content per cell, lysed cells were a major contributor of MCs to the extracellular environment, although the model overestimated the concentrations. Relationships for Planktothrix were significant but weaker, possibly due to reduced accuracy in the cell enumeration step, which would have altered the calculated MC content per cell. Whilst these findings support the hypothesis that cell lysis is the main contributor of extracellular MCs, the results do not exclude a role of MCs as signalling molecules. The recent finding that programmed cell death may occur in Microcystis under various environmental conditions may explain the commonly observed increase in extracellular MCs. Understanding the mechanisms involved in the transfer of MCs to the extracellular environment will provide further clarification on the function of these secondary metabolites and lead to the improvement of water quality management strategies.</p>

Investigating the Mechanism Behind the Release of Microcystins in Freshwater Cyanobacteria

<p>The frequency and distribution of toxic cyanobacterial blooms are increasing globally, creating the need for a better understanding of the processes involved in toxic secondary metabolite production. Microcystins (MCs) are potent hepatotoxins produced by a wide range of bloom-forming cyanobacteria genera such as Microcystis and Planktothrix. Although the release of MCs to the extracellular environment has long been considered a by-product of cell lysis and death, several studies suggest the presence of a mechanism that actively transports these toxins outside the cell membrane. The aim of the present study was to find evidence for a link between cell lysis and concentrations of extracellular MCs. A dual-fluorescence cell viability assay using the nucleic acid stain SYTOX Green was optimised for use on Microcystis and Planktothrix. A SYTOX Green concentration of 1 µM, and an incubation time of 30 minutes, yielded a bright and even fluorescent signal that readily identified lysed cells. The improved staining technique, in conjunction with liquid chromatography-mass spectrometry analyses, was employed in a culturing experiment to track the transfer of MCs to the extracellular environment in relation to the amount of cell lysis. For Microcystis, there was a strong and significant positive relationship between cell lysis and the concentration of extracellular MC. When the extracellular MC was predicted according to cell lysis levels and the MC content per cell, lysed cells were a major contributor of MCs to the extracellular environment, although the model overestimated the concentrations. Relationships for Planktothrix were significant but weaker, possibly due to reduced accuracy in the cell enumeration step, which would have altered the calculated MC content per cell. Whilst these findings support the hypothesis that cell lysis is the main contributor of extracellular MCs, the results do not exclude a role of MCs as signalling molecules. The recent finding that programmed cell death may occur in Microcystis under various environmental conditions may explain the commonly observed increase in extracellular MCs. Understanding the mechanisms involved in the transfer of MCs to the extracellular environment will provide further clarification on the function of these secondary metabolites and lead to the improvement of water quality management strategies.</p>

APLICACIÓN DE COLORACIONES ESPECIALES EN EL DIAGNÓSTICO HISTOPATOLÓGICO: TINCIÓN TRICRÓMICA DE MASSON

Masson's trichrome (MT) is a three-color staining protocol used in histology. MT allows to show and quantify changes such as tissue repair (healing) and collagen deposition. Also, it can be used to quantify blood vessels, in epithelial dysplasia and squamous cell carcinoma. The objective of this work is to describe the MT staining technique and to exemplify some applications of this technique in routine veterinary histopathological diagnosis. Archived histologic sections were selected from the records of the histopathology laboratory. Tissues were selected in base on theirs structures and lesions that could be evaluated with MT: a rabbit lung with a chronic suppurative bronchopneumonia; a bovine liver with lesions of Echium plantagineum poisoning; and a bovine eyelid with a squamous cell carcinoma. The TM was able to show fibroplasia in the pulmonary interstitium and confirm the presence of a chronic respiratory process, and clearly revealed an abundant fibrovascular tumor stroma, with profuse connective tissue and neovascularization between the tumor cells in deep dermis. In the liver, extensive and marked fibroplasia was confirmed. MT represents a complementary coloration to routine hematoxylin and eosin stain (H&E) and provides accurate information from several pathological processes, mainly those related to fibrovascular proliferation and scarring.

Effects of Thymoquinone on Adipocyte Differentiation in Human Adipose-Derived Stem Cells

Background: Obesity is one of the most important public health problems worldwide. Stem cells are primary cells capable of differentiating into different types of cells, and can be used to treat various diseases. Thymoquinone (TQ) has antioxidant, anti-inflammatory, anti-diabetic and anti-obesity properties. Herein, we aim to investigate the effect of TQ on the process of lipid differentiation in human adipose tissue-derived stem cells (ADSCs). Methods and Results: Quantification of cell surface markers was used by Flow-Cytometry and then, the effect of TQ on cell viability was assessed using alamarBlue test. ADSCs were then subjected to induction of differentiation in the presence of non-cytotoxic concentrations of TQ (6.25, 12.5 and 25 μg/mL). ADSCs differentiation was assessed using Oil-Red staining technique. Moreover, expression of PPARγ (Peroxisome proliferator activated receptor γ) and FAS (Fatty Acid Synthetase) proteins was evaluated using Western blotting analysis. Flow-cytometric analysis demonstrated the expression of CD44 and CD90 markers as mesenchymal stem cells markers on the surface of ADSCs. At concentrations≤100 μg/mL of TQ, no significant difference in cell viability of ADSCs was observed compared to the control. Adipocyte differentiation process significantly decreased at 25 μg/mL (P<0.001) and 12.5 μg/mL (P<0.01) of TQ. The results of the qualitative examination of Lipid Droplets also confirmed these results. Western-blot analysis showed that TQ at 12.5 (p<0.05) and 25 μg/mL (p<0.01) reduced FAS/β-actin ratio compared to the positive group.Conclusions: This study showed that TQ can reduce the process of differentiation of fat stem cells into fat cells and might be considered as an anti-obesity compound.

Von Kossa and his staining technique

One hundred and twenty years ago, the Hungarian physician Julius von Kossa developed a now classical staining method for detecting mineral deposits in animal tissues. Since then, this method has been widely adapted and combined with different counterstains, but still bears the name of its original inventor, who, if alive, would have turned 150 in 2015. As a rather inexpensive technique that does not require special instrumentation, von Kossa’s staining method became extremely popular for demonstrating mineralized tissues in vivo and in vitro. This article pays tribute to von Kossa and to his handy staining method.