EXAMINATION OF THE DENSITY OF SEMEN AND ANALYSIS OF SPERM CELL MOVEMENT. 1. INTRODUCTION

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1 JOURNAL OF MEDICAL INFORMATICS & TECHNOLOGIES Vol.3/00, ISSN Łukasz WITKOWSKI * mage enhancement, mage analyss, semen, sperm cell, cell moblty EXAMINATION OF THE DENSITY OF SEMEN AND ANALYSIS OF SPERM CELL MOVEMENT. The algorthms for examnaton of the densty and chosen parameters of movement of sperm cell were elaborated and mplemented. The conducted research s a part of a work on a computer system for analyss of semen. The system wll allow for an ncrease of the precseness of examnaton thanks to the exact specfcaton of numercal values of chosen parameters. Addtonal advantage of the system s a shorter tme of the examnaton. Nowadays, the basc type of examnaton s an estmated analyss of parameters of semen done by vsual observaton of a sample. Ths type of examnaton s based on a subjectve assessment of an mage by a physcan. Moreover the regstraton of mages n vsual analyss s not possble.. INTRODUCTION One of the more and more sgnfcant problems n the present world becomes the queston of fertlty of men. Researches conducted snce many years show that the percentage of men havng problems wth fertlty has been ncreasng. Ther semen s weak, characterzed by low densty, small fracton of sperm cells wth proper structure and movement. All these make them unable for natural fertlzaton. As a consequence of the development of the technques of n vtro fertlzaton, the assessment of the qualty of semen becomes vtal. In many cases thanks to these technques artfcal fertlzaton s possble despte of weak sperm. As a result of long year researches, World Health Organzaton standards have been elaborated. They classfy the qualty of semen on the bass of the value of such coeffcents as: densty, percentage of sperm cells wth proper structure, fracton of sperm cell wth proper movement etc. Healthy semen s characterzed by the densty of 0 50 mllons sperm cells n ml. as well as by the fracton of at least 5% of sperm cells wth proper.e. lnear and progressve movement. Problematc s, however, the way n whch these coeffcents can be determned. Currently the most common method used to fnd out the above parameters s vsual analyss carred out by andrologst. The analyss s a subjectve assessment of a mcroscope sample. The precseness of such examnaton could be questonable and t depends on the experence of doctor. The possblty of repetton of the examnaton does not exst. One cannot ether regster mages or create hstory of nvestgated samples. Ths problem s especally mportant n the case when a * Instytut Bocybernetyk Inżyner Bomedycznej, Polska Akadema Nauk, Warszawa ul. Trojdena 4, e-mal: lw@bb.waw.pl

2 patent changes hs andrologst. A new physcan has to examne hs patent once agan and the results of hs analyss may vary sgnfcantly from the ones done by the prevous physcan. The development of the computer technques caused an attempt for the elaboraton of such system that would elmnate the mprecseness of vsual analyss. Ths system would allow for determnaton of exact value of the coeffcents, for regstraton of mages and creaton of the hstory of examned samples. In 80 CASA (Computer Asssted Semen Analyss) systems were created. They were based on sgnal processors amed at fast processng of mages. Dsadvantage of these systems was ther hgh prce only few clncs could afford t. The requrement of easy access was not fulflled. Stll the man analyss was vsual examnaton. Thanks to the further development of computers and the decrease of ther prces t s easy to fulfll the postulate of accessblty. Therefore I decded to elaborate a computer system for examnaton of semen. It would requre a computer synchronzed wth a mcroscope. The frst stage of the work was to prepare an applcaton enumeratng the densty of sperm. The second step was creaton of an applcaton that would determne parameters of sperm movements.. MATERIALS Thanks to the co-operaton wth the st Obstetrc-Gynecologcal Clnc of professor Bablok n Warszawa t was possble to have an access to samples of semen of patents examned n the clnc. To elaborate and test algorthms a sequence of mages of semen of a healthy patent was regstered. It conssts of 50 subsequent monochromatc mages of sze 400 X 400 pxels and 56 levels of gray. The tme of a sequence s around 4 seconds (50 mages at Hz). Fg.. Subsequent mages of a regstered sequence (x0) The sample s a sequence of alve semen mages. On account of regstraton of alve sperm cells the semen s not staned. Therefore, the mages are not contrast and t s dffcult to extract sperm cells (one should extract sperm cells and reject any artfacts and other elements of semen that are not sperm cells before the analyss). MI - 8

3 3. IMAGE ANALYSIS 3.. PRE-PROCESSING In order to calculate semen coeffcents t s necessary to extract sperm cells from the background. To realze the task one uses operatons of mage enhancement lke: logcal, arthmetc, morphologc, neghborhood, pont to pont. Addtonal and mportant help s a possblty of detecton of movement on the two followng mages by calculaton of ther sum. Also avalable s further processng of the sum of the mages. After the set sequence of operatons has been done next step s the detecton of a gravty center of sperm cells on consecutve mages of the sample. 3.. TRAJECTORIES After the process of pre-processng and extractng of objects (sperm cells) t s possble to search trajectores of sperm cells movements by jonng ther gravty centers. Some knowledge of semen movement s known from the lterature. It s known that the maxmum speed of a sperm cell s 5 μm/s. It can be concluded from ths data, knowng that the mcroscope magnfcaton was 0 tmes and camera was capable of capturng mages per second that the gravty center of a sperm cell can change ts locaton maxmally by 0 pxels on a sngle consecutve mage. Another mportant feature of sperm cell movement s the ablty to move only forward. Those condtons n most cases make possble to choose accurately the next proper gravty center. In the case of conflct (more than one gravty center meet prevous condtons) to choose properly between possble trajectores one can take nto consderaton addtonal nformaton: speed. It s known that the speed of a sperm cell cannot change sgnfcantly. Ths allows for an accurate and approprate determnaton of the next gravty center for a gven trajectory DENSITY The number of sperm cells (l) s determned on the bass of the number of found trajectores of sperm cell movements, that had ether proper or mproper run, observed n a sequence of 50 mages. On the bass of the number of sperm cells one calculates the densty of the sample ( ρ sample ), where under the sample one understands the quantty of ejaculate used to prepare the mcroscope sample: ρ sample l v sample * S S o where: v sample l average number of sperm cells wthn the area of a sngle mage, precsely measured quantty of the semen used for preparaton of the mcroscope sample, S area of a mcroscope sample, S o area of a sngle mage MI - 83

4 The densty of the sample s calculated on the bass of average value of the number of the sperm cells wthn the area of a sngle mage, as the number of sperm cells s dfferent n the consecutve mages. On the bass of the densty of the sample one can calculate the number of sperm cells (L) n the sample of semen. L ρ * samlpe V ejaculate 3.4. THE PARAMETERS OF MOVEMENT. Another mportant parameter evaluatng the qualty of the semen s sperm cells movements. Accordng to WHO standards to defne the qualty of sperm cell movement (ndex ) one has to calculate the followng parameters of ts trajectory: VSL straght-lne velocty: VSL ( x M x ) + ( y M ( M ) Δt y ) where: ( x, y ) locaton of the gravty center of a sperm cell on the frst mage, ( x, y M M ) locaton of the gravty center of the sperm cell on the fnal (M) mage, Δt dfference of tme between the subsequent mages VCL curvlnear velocty: VCL M j ( x j+ x ) j + ( y ( M ) Δt j+ y ) j VAP average path velocty: VAP M j ( x j+ x j ) + ( y ( M ) Δt j+ y j ) + k k x 5 k k where: x and + y k y respectvely 5 k STR straghtness: devaton of the averaged route n relaton to the straght lne: VSL STR VAP LIN lnearty: devaton of the trajectory n relaton to the straght lne: MI - 84

5 VSL LIN VCL Fg.. Illustrated parameters of the sperm cell movement 4. IMPLEMENTATION OF SEMEN ANALYSIS ALGORITHMS. Worker out applcaton realzes the followng tasks:. enhancement of mages n a such way that one could extract objects from the background. rangng out trajectory of the movement of sperm cell by jonng gravty centers of extracted objects 3. calculaton of the parameters of movement for each trajectory 4. calculaton of the densty of a sample 4.. PRE-PROCESSING The followng set of operatons realzes the frst of tasks, that s: enhancement of mages: medan flterng. multplyng and scalng. left shft (scalng) blur negaton Max flter - flterng gvng the max value from the neghborhood Those operatons are performed on two subsequent mages. Then the mages are subtracted one from the other and the followng operatons are processed: square dlaton closng eroson thresholdng dlaton eroson MI - 85

6 Fg.3. Wndow for defnng of the sequence of operatons 4.. TRAJECTORIES The worked out applcaton extracted objects on bass of threshold operaton. Then the gravty centers were found for all the extracted objects and generated junctons between found centers of gravty traced trajectores. There were trajectores n the sample: 0 of them of proper run and of a bad run. A problem of napproprate sperm cell movement was met. In two cases only the frst condton (max. 0 pxel replacement) was fulflled. The second (forward movement) was not. To solve the problem n the frst processng several separate trajectores were found. After that n the second step those peces were joned nto a sngle trajectory of bad sperm cell run. The second step dd not make any nfluence on trajectores of well buld sperm cells. Fg.4. Trajectores MI - 86

7 4.3. DENSITY The applcaton calculated the densty of the semen on the bass of quantty of trajectores n 50 mages. Ths s only a plot evaluaton of the semen. In the future the densty of a sample should be evaluated on the bass of bgger amount of mages of the same patent THE PARAMETERS OF MOVEMENT. The calculated values of movement confrmed a pror observaton of trajectores. 0 of them were proper and were wrong. The table below presents the result of measurements of all the parameters of cells movements for trajectores. Those marked ndcated mproper trajectores. Fg.5. Trajectores Accordng to WHO standards fracton of at least 5% of proper movement sperm cells classfes patent as healthy. In the examnaton of our patent only 7 % of all trajectores were mproper. It confrmed the patent was healthy. 5. CONCLUSION The mplemented applcaton allowed for an exact determnaton of the parameters of sperm cells movement. However, further research on clncal materal s requred to verfy approprateness of the mplemented. Further researches are needed also to calculate the densty of sperm n order to answer the queston how bg should be the sequence of mages to determne densty of semen wth an assumed precson. It seems that the worked out applcaton performs ts tasks: t analyzes sample, provdes physcan wth substantal data, thanks to whch he can ssue more objectve dagnoss. A long year experence for a physcan would not be necessary. Easy way of handlng and accessblty of the MI - 87

8 most mportant components: computer and mcroscope would allow a broad applcaton of the system n many medcal centers n the future. The applcaton was developed n Pascal language. The tools of the Borland frm (Delph verson 5) and lbrary: Intel Image Processng Lbrary verson.5 was used. The applcaton s compatble wth Wndows NT 4.0/000. The tests were performed on computer PC wth a processor Pentum III 450 MHz. BIBLIOGRAPHY [] GONZALES R. C., WINTZ P., Dgtal Image Processng, Addson-Wesley, 987 [] NADLER M., SMITH E., Pattern recognton engneerng, John Wley & Sons, 99 [3] Intel Image Processng Lbrary, reference manual, Intel Corporaton, 000 [4] Techncal Gude for IVOX, TOX IVOS, CEROS, Hamlton Thorne Research, 000 [5] SEMCZUK M., KURPISZ M., Androloga, PZWL, 998 [6] RADWAN J., Kompleksowa ocena wybranych typów spermogramów spermocytogramów jej wartość klnczna prognostyczna, praca habltacyjna, Akadema Medyczna w Łodz, Łódź, 994. [7] World Health Organzaton, WHO Laboratory manual for the Standardzed Investgaton and Dagnoss of the Infertle Couple, Cambrdge Unversty Press, 993 MI - 88