infertility and sperm analysis
DESCRIPTION
Spermatozoon,Male Infertility,Female infertility,Tests and Diagnosis,Hydrosalpinx,Fern test,Semen analysis,IVFTRANSCRIPT
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INFERTILITY AND SEMEN ANALYSIS
Dr VikashJR(Pathology)
IMS,BHU
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Infertility-Apparent faliure of a couple to conceive
Sterility-Absolute inability to conceive.
If a couple fails to achieve pregnancy after 1 year of unprotected and regular intercourse , it is an indication to investigate the couple
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Type
Primary- If conception has never occurred. Secondary
It may be physiological
Before puberty and after menopause
Over age of 40 year fertility reduced and increase risk for chromosomally and other malformed foetus.
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Symptoms
Woman: abnormal menstrual periods Man: hormonal problems (changes in hair
growth or sexual function)
Infertility is rarely absolute so the term sub-fertility may be more appropriate
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Spermatogonia--Mitosis--spermatocytes—Meiosis I-Haploid Secondary Spermatocytes---Meiosis II-Spermatid ( 74 days) Spermiogenesis:
differentiation of the round spermatid into a spermatozoon
This is the process in which sperm morphology is largely determined
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What the spermatozoon looks like
The human sperm cell is about 70 µm long
The nucleus is in the head – contains the 23 chromosomes
It is the head which binds to the egg at fertilization
Midpiece: the energy for motility is generated
Tail: motility – the beat is initiated just behind the midpiece, and then propagated along the tail
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BACKGROUND INFORMATION
At puberty there are 300,000 primordial follicles
Dominant follicle produces oestradiol which leads to LH surge
Ovulation occurs 24-36 hours later The fertilization life span of the ovum is
24-36 hours The receptivity of the endometrium is
days 16-19 of a 28 day cycle
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Hypothalmic-Pitutary-Gonadal Axis
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Causes of male infertility
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Normal Semniferous tubule
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Tubular atrophy
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Causes of female infertility
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Causes of female infertility Dyspareunia and Vaginal Causes Congenital Defect in genital tract-Absent or septate
vagina,Hypoplasia Infection- Chlamydia Cervicitis Cervical factor-cervical mucous Uterine causes-Hypoplasia,Malformed uterus and
incomplete os Tubal factor-Salpingitis,Gonorrhoea ir chlamydial
infection Ovaries-PCOD,LFD Peritoneal causes-Pelvic endometriosis Chronically ill health Hormonal pitutary dysfuction-
Hyperprolactinoma,Hypothalmic disease
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Risk factors
Age Tobacco smoking Alcohol use Being overweight Too much exercise Caffeine intake
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Tests and diagnosisTests for men General physical examination Semen analysis Hormone testing Transrectal and scrotal ultrasoundTests for women Ovulation testing Hysterosalpingography Laparoscopy Hormone testing Ovarian reserve testing Genetic testing Pelvic ultrasound
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Tests History Examination Specific test- Hysterosalpingography(HSG) Laproscopic chromotubationSonosalpingography(SSG)Hysteroscopy and falloscopyAmpullary and fimbrial salpingoscopyEndometrial BiopsyFern TestUltrasoundHormonal Test
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Hysterosalpingography(HSG)
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Hydrosalpinx
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FERN Test
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Semen
A mixture of seminal plasma and cells Seminal plasma contains:
Prostatic fluid (~30% of the volume) Epididymal plasma (~5% of the volume) Seminal vesicle fluid (the remainder of the ejaculate)
The cells are: Spermatozoa Germ line cells Leukocytes of various types Bacteria Epithelial cells Occasional red cells
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Semen Analysis
• There are several macroscopic evaluations which give useful diagnostic information about the sample:
–Appearance
–Odour
–Liquefaction
–Volume
–Viscosity
–pH
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SEMEN ANALYSIS• pH is important because sperm die at pH < 6.9
• The pH of liquefied semen is normally determined using test strips (we use EM Science ColorpHast type, pH 6.5–10.0)
• We usually measure pH after volume and viscosity – by touching the “emptied” volumetric pipette to the test strip
• The normal pH range is 7.2–8.4• Inflammatory disorders of the accessory glands can take the pH outside of this range
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Semen Analysis-Assesement
• The characteristics assessed are:– Motility– Sperm aggregation (random clumping) – “some” is normal,
but large clumps (each with hundreds of sperm) is abnormal
– Spermagglutination (between specific sites) – could suggest the presence of antisperm antibodies.
– Round cells: should be <1 per 40× field (~ 1 million/ml). If more abundant, a leukocyte test should be run
– Epithelial cells: usually present in small numbers– Erythrocytes: should not be present– Debris: particles smaller than sperm head, may be plentiful– Bacteria and protozoa: presence indicates infection
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Semen Analysis-Motility Assessment
• % motile = the proportion of sperm with tail movement
• Progression rating = the grade of progression shown by the majority of the sperm: this can be from 0 (all immotile) to 4 (all with rapid progression); or from a (rapid progression) to d (all immotile)
• Differential motility count = proportion of sperm in each of 4 motility classes (rapid progressive; slow progressive; non-progressive; immotile
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Motility Asses. Contd.
• Differential motility classification is based on the distance swum over time:
– Rapid progressive: > 25 µm/s
– Slow progressive: 5 – 25 µm/s
– Non-progressive: < 5 µm/s
– Immotile: no flagellar movement
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Sperm Morphology Morphology is even more important than
motility and concentration Because of the small size of the human sperm
head, must use an air-dried smear which has been stained
The Papanicolaou method is best Prepared samples are assessed using a 100×
oil-immersion objective under bright field optics The WHO recommends that 200 spermatozoa
are counted per sample (and says that 2 × 200 is better)
Fields for counting must be selected at random When counting, remember about the normal
distribution
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Morph. Contd.
Variations of norm al head shape
Sm all / large head Tapering heads
Pyriform heads Vacuolatedhead
Asym m etricinsertion
D istendedm idpiece
Thinm idpiece
C ytoplasm icdroplet
C oiledta il
Shortta il
H airp inta il
Bentta il
Term inal droplet
D uplicateta il
C onjo inedform
N on-insertedta il
Am orphous form s
C onstricted R educedacrosom e
D ensesta in ing
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Sperm morphology – TZI The Teratozoospermic Index is an expression of the
average number of abnormalities per abnormal sperm Each sperm cell is assessed for an abnormality in the
head, neck/midpiece, or tail, and for a cytoplasmic droplet
If it does not have any of these abnormalities, it is “normal”
If it does have an abnormality, it is “abnormal”, and we score each abnormality. So, if a cell has an abnormal head and tail, it is counted as 1 cell, and 2 abnormalities
Then, (total # abnormalities) / (total # sperm) = TZI A TZI > 1.80 has been associated with poor sperm
fertilizing ability in vivo and in vitro
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Semen Biochemistry Acid phosphatase: marker for prostatic function
Citric acid: can indicate prostatic function – low levels may indicate dysfunction or a prostatic duct obstruction
Zinc: marker for prostatic function – colorimetric assay (WHO)
Fructose: marker for seminal vesicle function, and is a substrate for sperm metabolism – spectrophotometric assay (WHO)
-Glucosidase: secreted exclusively by the epididymis and so is a marker for epididymal function – spectrophotometric assay (WHO)
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Terminology
Aspermia-No semen Azoospermia-No sperm in semen Oligospermia-Low sperm count Asthenospermia-Dimnished Motility Necrospermia-Dead sperm Teratospermia-Abnormal Morphology
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FERTILITY TREATMENT OPTIONS
If a partner is sterile (i.e. no gametes), then the couple would need donor gametes to achieve a pregnancy
If one or both partners are sub-fertile, then the treatment options are:
no treatment, or ovulation induction
intra-uterine insemination (+ ovulation induction)
in vitro fertilization (includes ICSI)
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INTRA-UTERINE INSEMINATION (IUI)
IUI is the least invasive of all t/t - involves the selective washing of semen to isolate the motile spermatozoa (can’t put whole semen into the uterus)
Up to 15 million motile spermatozoa are inseminated
Advantages: relatively inexpensive – simple procedures minimal use of FSH can be used in consecutive cycles can usually start treatment virtually immediately
Disadvantages: lower success rate per cycle than other treatment
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IN VITRO FERTILIZATION (IVF)Overview
There are many types of IVF
For virtually all types, the woman is treated with “fertility drugs” to stimulate the development of a group of eggs (the average is around 10 – but the range can be enormous)
Just prior to ovulation, the oocytes are retrieved
That afternoon, they are inseminated with prepared sperm
Inseminated eggs checked the next day for fertilization
The fertilized eggs are kept in culture for up to 5-6 days
Embryo transfer / possibly cryopreservation
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IVF-Embryo development
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IVF Contd.
One sperm is injected directly into an egg
Only mature eggs injected
After the insemination, the rest of the lab procedures are the same as for “standard” IVF
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Thank you