Morphometric characterization and classification of vicuña sperm (Vicugna vicugna) using ISAS® CASA-Morph system

The aim of the study was to determine the morphological and morphometric characteristics of the vicuña sperm (Vicugna vicugna), looking for possible subpopulations in its structure. Semen was collected by electroejaculation method from three adult male vicuñas weighing 50.33 ± 2.52 kg, once per animal. The volume, color and concentration, sperm morphology and morphometry were determined. The samples for the analysis of sperm morphology and morphometry were stained with Hemacolor®, and analyzed using the CASA-Morph, Integrated Semen Analysis System (ISAS®v1). Five forms of the vicuña sperm head were distinguished: normal, piriform, long, short and rounded. The morphometric parameters determined were the length, width, area, perimeter, ellipticity, elongation, regularity and rugosity of the vicuña sperm head, percentage of acrosome, head gray level, as well as the width, area, distance and angle of insertion of midpiece of the sperm. In relation to morphology, significant differences (P <0.05) were found in the percentage distribution of head shapes, with the normal shape (55.7%) being the majority and different from the other shapes. Significant differences (P <0.05) were found between animals in the morphometric variables of head, percentage of acrosome, ellipticity, rugosity, elongation and gray level; while the variables of the midpiece and regularity did not show differences (P> 0.05). The morphometric variables were distributed in four main components (PCA) called elongation, area, circularity and midpiece width, which explained 84.59% of the total variance. The cluster analysis determined five subpopulations (SP): SP1 grouped small cells of low length, width and area (18.8%); SP2 of sperm of large size, both in area and width of the head (17.38%); SP3 of rounded cells with high values of percentage of acrosome and head gray level (24.04%); SP4 of spermatozoa of intermediate size and elongated, with greater ellipticities and elongation (23.61%); and SP5 of cells of intermediate size and short, with low values of area and length (16.71%).

Summary of four scientific studies on Arsenicum album high dilution effect against Arsenic intoxication in mice

Background: Groundwater arsenic affects millions of people in about 20 countries. In West Bengal (India) and Bangladesh alone over 100 million people are exposed. The arsenic concentration in contaminated groundwater in Bangladesh was above the maximum permissible level of 0.05 mg/l as recommended by WHO for developing countries [1]. Drinking water is not the only source of poisoning. In arsenic contaminated areas, crops, vegetables, cereals, poultry, cattle, etc, also contain traces of arsenic. Chronic arsenic intoxication has been associated with several diseases such as melanosis, leuco-melanosis, hyperkeratosis, oedema, skin cancer… Cazin et al [2], have demonstrated the effect of high dilutions of arsenic compounds. They noted increased arsenic elimination from blood through urine and faeces in intoxicated rats. According to these research, the aim of Khuda Buksh studies [3-4-5] was to investigate whether high dilution Arsenicum album have any effect on arsenic accumulation in different tissues and to understand also how this high dilution could produce a protective effect on all the different organs. Methodology: Firstly, the effect of Arsenicum album 30 cH on the amount of arsenic accumulation was determined by spectrophotometric analysis in four tissues namely liver, kidney and testis in mice intoxicated by arsenic. The protective effect in chronic and acute arsenic intoxicated mice of Arsenicum Album 6cH, 30cH and 200cH has been evaluated using not only the activities of enzymatic and biomarker toxicity (aspartate amino transferase (AST), alanine amino transferase (ALT), acid phosphatase (AcP), alkaline phosphatase (AlkP), lipid peroxidation (LPO) and reduced glutathione (GSH)) but also the cytogenetical parameters (chromosome aberrations (CA), mitotic index (MI), sperm head anomaly (SHA) etc., ). Because, it is well demonstrated that these enzymes biomarkers reflect the degree of hepatotoxicity and oxidative stress caused by arsenic intoxication. Results: Compared to controls, Arsenicum album 30cH induced a significant decrease in accumulation of arsenic in 4 tissues namely liver, spleen, kidney and testis in intoxicated mice. In addition, both Arsenicum album 6cH, 30cH and 200cH reduced chromosome aberrations, sperm head abnormality frequencies and activities of acid and alkaline phosphatases, aspartate and alanine aminotransferases and lipid peroxidation, while mitotic index and activities of glutathione, catalase and succinate dehydrogenase were increased compared to controls. Conclusion: Altogether, theses results provide evidence of protective potentials of the Arsenicum album dilution against acute and chronic arsenic intoxication in mice. They also offer a new hypothesis that the mechanism of the homeopathic dilution could act through regulation of expression of certain genes. This explanation seems to be plausible because all biomarker tests are regulated by specific genetic regulatory mechanisms [6]. keywords: Arsenicum album, arsenic intoxication, enzymatic and biomarker toxicity. References: [1] WHO. WHO Guidelines for Drinking Water Quality, Vol. 2, 2nd edition. Geneva: WHO, 1996, 940–9. [2] Cazin JC, Cazin M, Gaborit JL, Chaoui A, Boiron J, Belon P, et al. A study of the effect of decimal and centesimal dilutions of arsenic on the retention and mobilization of arsenic in the rat. Hum Toxicol 1987;6:315–20. [3] Mitra K, Kundu SN, Khuda-Bukhsh AR. Efficacy of a potentized homoeopathic drug (Arsenicum Album-30) in reducing toxic effects produced by of arsenic trioxide in mice. I. On rate accumulation of arsenic in certain vital organs. Comp Ther Med 1998;6:178–84. [4] Pathikrit Banerjee. Evidences of Protective Potentials of Microdoses of Ultra-high Diluted Arsenic Trioxide in Mice Receiving Repeated Injections of Arsenic Trioxide. eCAM 2009; 1-10. [5] Pathikrit Banerjee, Comparative Efficacy of Two Microdoses of a Potentized Homeopathic Drug, Arsenicum Album, to Ameliorate Toxicity Induced by Repeated Sublethal Injections of Arsenic Trioxide in Mice. Pathobiology 2008;75:156–170. [6]/ Khuda-Bukhsh AR. Potentized homeopathic drugs act through regulation of gene expression: a hypothesis to explain their mechanism and pathways of action in vivo. Comp Ther Med 1997;5:43–6

Local Constraint and Label Embedding Multi-layer Dictionary Learning for Sperm Head Classification

Morphological classification of human sperm heads is a key technology for diagnosing male infertility. Due to its sparse representation and learning capability, dictionary learning has shown remarkable performance in human sperm head classification. To promote the discriminability of the classification model, a novel local constraint and label embedding multi-layer dictionary learning model called LCLM-MDL is proposed in this study. Based on the multi-layer dictionary learning framework, two dictionaries are built on the basis of Laplacian regularized constraint and label embedding term in each layer, and the two dictionaries are approximated to each other as much as possible, so as to well exploit the nonlinear structure and discriminability features of the morphology of human sperm heads. In addition, to promote the robustness of the model, the asymmetric Huber loss is adopted in the last layer of LCLM-MDL, which approximates the misclassification error by using the absolute error function. Finally, the experimental results on HuSHeM dataset demonstrate the validity of the LCLM-MDL.

Bayesian Pixel Density Estimation Modeling to Detect Human Sperm Sample Image Based on Sperm Head Shape

Currently, there is a problem of the difficulty in classifying human sperm head sample images using different databases and measuring the accuracy of several different datasets. This study proposes a Bayesian Density Estimation-based model for detecting human sperm heads with four classification labels, namely, normal, tapered, pyriform, and small or amorphous. This model was applied to three kinds of datasets to detect the level of pixel density in images containing normal human sperm head samples. Experimental results and computational accuracy are also presented. As a method, this study labeled each human sperm head based on three shape descriptors using the formulas of Hu moment, Zernike moment, and Fourier descriptor. Each descriptor was also tested in the experiment. There was an increased accuracy that reached 90% after the model was applied to the three datasets. The Bayesian Density Estimation model could classify images containing human sperm head samples. The correct classification level was obtained when the human sperm head was detected by combining Bayesian + Hu moment with an accuracy rate of up to 90% which could detect normal human sperm heads. It is concluded that the proposed model can detect and classify images containing human sperm head objects. This model can increase accuracy, so it is very appropriate to be applied in the medical field

The social shape of sperm: using an integrative machine-learning approach to examine sperm ultrastructure and collective motility

Sperm is one of the most morphologically diverse cell types in nature, yet they also exhibit remarkable behavioural variation, including the formation of collective groups of cells that swim together for motility or transport through the female reproductive tract. Here, we take advantage of natural variation in sperm traits observed across Peromyscus mice to test the hypothesis that the morphology of the sperm head influences their sperm aggregation behaviour. Using both manual and automated morphometric approaches to quantify their complex shapes, and then statistical modelling and machine learning to analyse their features, we show that the aspect ratio of the sperm head is the most distinguishing morphological trait and statistically associates with collective sperm movements obtained from in vitro observations. We then successfully use neural network analysis to predict the size of sperm aggregates from sperm head morphology and show that species with relatively wider sperm heads form larger aggregates, which is consistent with the theoretical prediction that an adhesive region around the equatorial region of the sperm head mediates these unique gametic interactions. Together these findings advance our understanding of how even subtle variation in sperm design can drive differences in sperm function and performance.

Genotoxic activity of betel nut on germinal cell in Sarcoma 180 ascites tumour bearing male mice

The genotoxicity of the ethanolic extract of betel nut was evaluated using sarcoma 180 tumour bearing mouse considering sperm motility, sperm viability, biochemical estimation of fructose in seminal fluid and sperm head morphology assays. Sperm head morphology was studied by H-E staining and Toluidine blue staining method. But Toluidine blue staining method is a reliable method to evaluate the DNA damage of sperms. Ethanolic BNE (betel nut extract) can suppress the percentage of sperm motility, sperm viability and seminal fructose level. In addition, it can also enhance the percentage of DNA damaged sperms. Moreover, histological sections of testes have been studied in control and BNE treated sarcoma 180 tumour bearing mice to highlight the potential toxic effect of BNE. The significant decreasing rate of seminal fructose concentration, sperm motility as well as viability and increasing rate of sperm head abnormality in different doses of treated series may be as a result of different toxic alkaloid ingredients present in BNE. Therefore, the results showed the potential of the BNE to induce different types of germ cell abnormalities in tumour bearing male mice.