4. Morphological abnormalities of spermatozoa

The abnormalities in morphology of spermatozoa have a negative effect on the outcome of IVF and ICSI. Morphologically abnormal spermatozoa analyzed by light microscopy or MSOME are categorized into subgroups according to the defects of the head, neck, midpiece and/or tail.

#### 4.1. Abnormalities of the head

Spermatozoa, with head size and shape abnormalities (large, small, tapered, pyriform, round and amorphous heads), vacuolated heads, heads with small acrosomal area (<40% of head area), with double or multiple heads and without head are often found in the semen samples (Figure 3).

condensed chromatin, diploidy and aneuploidy of spermatozoa. Detailed genetic analysis shows that large-headed spermatozoa of some patients can have the homozygous mutation (c.144delC) in the Aurora kinase C (AURKC) gene. These spermatozoa of AURKC-deficient patients cannot complete meiosis in the absence of functional AURKC. Therefore they can be

Assessment of Human Sperm Cells Morphological Parameters

http://dx.doi.org/10.5772/intechopen.71413

57

Small-headed (microcephalous) spermatozoa are defined as those with a length < 3.5 μm and a width < 2.5 μm. These spermatozoa can have the excessive shrunk nucleus and abnormally condensed chromatin and fragmented DNA. They may also present with very small, abnor-

Elongated-headed spermatozoa are described as being a head length > 5 μm with a width < 3 μm or a length of <5 μm and a width of <2 μm. Pyriform heads are also included under elongation of spermatozoa. Elongation of head is generally recognized as a stressinduced sperm morphology aberration, and is prevalent especially in male accessory gland infections and in the presence of a varicocele. An abnormally elongated nucleus shape of these spermatozoa is related with defects in the nuclear membrane, an increased frequency of chromosomal aneuploidies and altered chromatin compaction. Also anomalies of the neck region, persistence of cytoplasmic residues can often exist in these spermatozoa [52, 53].

Globozoospermia (round-headed sperm syndrome) is genetically determined abnormalities of spermatozoa. This disorder is classified into two categories: (1) total and (2) partial globozoospermia. In the total globozoospermia, the spermatozoa are easily recognizable by their small, round head shape, high DNA fragmentation and the absence of the acrosome. Owing to the absence of acrosomes, the spermatozoa do not contain acrosomal enzymes. Therefore they are unable to bind and penetrate the zona pellucida and fuse with the oolemma of the oocyte. In the case of the partial globozoospermia, spermatozoa show the oval head shape with less condensed chromatin, a partially present or remnants of acrosome and a disorganized midpiece. Mitochondria are present not only in the midpiece, but in the spermatozoa head too [52, 54–56]. Amorphous-headed spermatozoa indicate the chromosome 18 disomy and sex chromosome aneuploidy. Also, the high incidence of amorphous heads is relative to other abnormal head

Multiple-headed spermatozoa have two or more closed or dissociated heads with or without an acrosome or midpiece. Disorders of movement and fragmented DNA are appropriated for

Human spermatozoa heads often contain vacuoles. They are not visible on standard magnification and can be observed using MSOME method only. According to their size, vacuoles are divided into three types: (1) large (>50% of surface area), (2) medium (50–25% of surface area) and (3) small (<25% of surface area of total sperm head). Based on the location, they are classified as acrosomal, equatorial and post-acrosomal. Also according to the size and the number of vacuoles in the head of one spermatozoon, they can be classified into three categories: (1) one small vacuole, (2) multiple sporadic vacuoles of various sizes and (3) one large vacuole. Head vacuoles are non-acrosomal origin large nuclear indentations of various sizes and positions, packed with membranous material organized in membrane whorls. Vacuoles

tetraploid [47–50].

forms [57].

these spermatozoa [51].

mally formed acrosomes [51, 52].

The cause of acephalic spermatozoa can be biallelic SUN5 mutations or homozygous deletion of SUN5 or mutation of BRDT and other genes [44–46].

Large-headed (macrocephalous) spermatozoa are defined as those with a length > 4.7 μm and a width > 3.2 μm. Large head can indicate insufficient shrinking of the nucleus, abnormally

Figure 3. Spermatozoa with abnormal head morphology. (a) Round head; (b) amorphous head; (c) large head and bent neck; (d) small tapered head and bent neck; (e) elongated head; (f) two-headed; (g) multiple-headed and (h) acephalic spermatozoon (arrow). Bar = 10 μm.

condensed chromatin, diploidy and aneuploidy of spermatozoa. Detailed genetic analysis shows that large-headed spermatozoa of some patients can have the homozygous mutation (c.144delC) in the Aurora kinase C (AURKC) gene. These spermatozoa of AURKC-deficient patients cannot complete meiosis in the absence of functional AURKC. Therefore they can be tetraploid [47–50].

4.1. Abnormalities of the head

56 Spermatozoa - Facts and Perspectives

spermatozoon (arrow). Bar = 10 μm.

of SUN5 or mutation of BRDT and other genes [44–46].

Spermatozoa, with head size and shape abnormalities (large, small, tapered, pyriform, round and amorphous heads), vacuolated heads, heads with small acrosomal area (<40% of head area), with double or multiple heads and without head are often found in the semen samples (Figure 3).

The cause of acephalic spermatozoa can be biallelic SUN5 mutations or homozygous deletion

Large-headed (macrocephalous) spermatozoa are defined as those with a length > 4.7 μm and a width > 3.2 μm. Large head can indicate insufficient shrinking of the nucleus, abnormally

Figure 3. Spermatozoa with abnormal head morphology. (a) Round head; (b) amorphous head; (c) large head and bent neck; (d) small tapered head and bent neck; (e) elongated head; (f) two-headed; (g) multiple-headed and (h) acephalic Small-headed (microcephalous) spermatozoa are defined as those with a length < 3.5 μm and a width < 2.5 μm. These spermatozoa can have the excessive shrunk nucleus and abnormally condensed chromatin and fragmented DNA. They may also present with very small, abnormally formed acrosomes [51, 52].

Elongated-headed spermatozoa are described as being a head length > 5 μm with a width < 3 μm or a length of <5 μm and a width of <2 μm. Pyriform heads are also included under elongation of spermatozoa. Elongation of head is generally recognized as a stressinduced sperm morphology aberration, and is prevalent especially in male accessory gland infections and in the presence of a varicocele. An abnormally elongated nucleus shape of these spermatozoa is related with defects in the nuclear membrane, an increased frequency of chromosomal aneuploidies and altered chromatin compaction. Also anomalies of the neck region, persistence of cytoplasmic residues can often exist in these spermatozoa [52, 53].

Globozoospermia (round-headed sperm syndrome) is genetically determined abnormalities of spermatozoa. This disorder is classified into two categories: (1) total and (2) partial globozoospermia. In the total globozoospermia, the spermatozoa are easily recognizable by their small, round head shape, high DNA fragmentation and the absence of the acrosome. Owing to the absence of acrosomes, the spermatozoa do not contain acrosomal enzymes. Therefore they are unable to bind and penetrate the zona pellucida and fuse with the oolemma of the oocyte. In the case of the partial globozoospermia, spermatozoa show the oval head shape with less condensed chromatin, a partially present or remnants of acrosome and a disorganized midpiece. Mitochondria are present not only in the midpiece, but in the spermatozoa head too [52, 54–56].

Amorphous-headed spermatozoa indicate the chromosome 18 disomy and sex chromosome aneuploidy. Also, the high incidence of amorphous heads is relative to other abnormal head forms [57].

Multiple-headed spermatozoa have two or more closed or dissociated heads with or without an acrosome or midpiece. Disorders of movement and fragmented DNA are appropriated for these spermatozoa [51].

Human spermatozoa heads often contain vacuoles. They are not visible on standard magnification and can be observed using MSOME method only. According to their size, vacuoles are divided into three types: (1) large (>50% of surface area), (2) medium (50–25% of surface area) and (3) small (<25% of surface area of total sperm head). Based on the location, they are classified as acrosomal, equatorial and post-acrosomal. Also according to the size and the number of vacuoles in the head of one spermatozoon, they can be classified into three categories: (1) one small vacuole, (2) multiple sporadic vacuoles of various sizes and (3) one large vacuole. Head vacuoles are non-acrosomal origin large nuclear indentations of various sizes and positions, packed with membranous material organized in membrane whorls. Vacuoles cannot be considered as degenerative structures but can be regarded as a normal feature of the sperm head and does not affect ICSI outcomes. Therefore, some vacuoles can include vacant or low density DNA or show DNA fragmentation or abnormal chromatin compaction. Large vacuoles in the nucleus can indicate aneuploidy, fragmented DNA and chromatin condensation defects in the cell [24, 55, 58–63].

All abnormalities of spermatozoa neck and tail reduce their motility and therefore have the

Assessment of Human Sperm Cells Morphological Parameters

http://dx.doi.org/10.5772/intechopen.71413

59

Bent necks are associated with DNA fragmentation. Abnormal tails (two-tailed, bent tails, irregular tails) are associated to chromosome 13 disomy, supernumerary chromosomal abnor-

The absence of a tail, the short and broken tail may be related to axonemal abnormalities that are only visible with electron microscopy, and an unknown genetic origin of these abnormalities could be hypothesized [64, 65]. Coiled tails of spermatozoa can be associated with varico-

The spermatozoa can often have cytoplasmic droplets and cytoplasmic residues. Cytoplasmic droplets are small, regular osmotically sensitive vesicles which are located at the neck as opposed to the end of the annulus (Figure 5). Cytoplasmic droplets of normal human spermatozoa are still present after ejaculation and have no negative influence on cell function.

Figure 5. Cytoplasmic droplets and residual cytoplasm structure ([68], open access). (A) Spermatozoa with cytoplasmic

malities and the cytoskeletal abnormalities (including centriolar defects) [57].

negative influence on fertilization of oocyte.

cele and epididymal dysfunction [66, 67].

droplets and (B) spermatozoa with residual cytoplasm.

#### 4.2. Abnormalities of the tail

Spermatozoa without tail or with the neck and midpiece defects (bent neck, asymmetrical insertion of the midpiece into the head, thick or irregular midpiece and abnormally thin midpiece) and principal and terminal pieces of tail defects (short, multiple, hairpin, broken, coiled, bent tail and tails of irregular width) are often found in the semen samples (Figure 4).

Figure 4. Spermatozoa with abnormal neck and tail morphology. (a) Absent tail; (b) coiled tail; (c) bent neck and (d) twotailed spermatozoon. Bar = 10 μm.

All abnormalities of spermatozoa neck and tail reduce their motility and therefore have the negative influence on fertilization of oocyte.

cannot be considered as degenerative structures but can be regarded as a normal feature of the sperm head and does not affect ICSI outcomes. Therefore, some vacuoles can include vacant or low density DNA or show DNA fragmentation or abnormal chromatin compaction. Large vacuoles in the nucleus can indicate aneuploidy, fragmented DNA and chromatin condensa-

Spermatozoa without tail or with the neck and midpiece defects (bent neck, asymmetrical insertion of the midpiece into the head, thick or irregular midpiece and abnormally thin midpiece) and principal and terminal pieces of tail defects (short, multiple, hairpin, broken, coiled, bent tail and tails of irregular width) are often found in the semen samples (Figure 4).

Figure 4. Spermatozoa with abnormal neck and tail morphology. (a) Absent tail; (b) coiled tail; (c) bent neck and (d) two-

tion defects in the cell [24, 55, 58–63].

4.2. Abnormalities of the tail

58 Spermatozoa - Facts and Perspectives

tailed spermatozoon. Bar = 10 μm.

Bent necks are associated with DNA fragmentation. Abnormal tails (two-tailed, bent tails, irregular tails) are associated to chromosome 13 disomy, supernumerary chromosomal abnormalities and the cytoskeletal abnormalities (including centriolar defects) [57].

The absence of a tail, the short and broken tail may be related to axonemal abnormalities that are only visible with electron microscopy, and an unknown genetic origin of these abnormalities could be hypothesized [64, 65]. Coiled tails of spermatozoa can be associated with varicocele and epididymal dysfunction [66, 67].

The spermatozoa can often have cytoplasmic droplets and cytoplasmic residues. Cytoplasmic droplets are small, regular osmotically sensitive vesicles which are located at the neck as opposed to the end of the annulus (Figure 5). Cytoplasmic droplets of normal human spermatozoa are still present after ejaculation and have no negative influence on cell function.

Figure 5. Cytoplasmic droplets and residual cytoplasm structure ([68], open access). (A) Spermatozoa with cytoplasmic droplets and (B) spermatozoa with residual cytoplasm.

Cytoplasmic residues are large, irregular material along the midpiece and can indicate the abnormal spermiogenesis (Figure 6). They form as outcome of incomplete cytoplasmic extrusion during spermiogenesis in the causes of suppression of FSH and/or androgens, deficiency of cyclin-dependent kinase 16 (CDK16) and organophosphorus pesticide (OP) exposure. In comparison to the cytoplasmic droplet, cytoplasmic residues contain major quantity of cytoplasm enzymes, which produce pathological amounts of reactive oxygen species. These species can negatively affect spermatozoa function including peroxidative damage to the cell membrane, DNA damage, mitochondrial dysfunction, impaired interaction with the female reproductive tract and lead to male infertility [1, 32, 52, 68, 69].

Multiple morphological abnormalities in the same patient's spermatozoa tail structure and ultrastructure which impair motility can show the mutation of some genes (DNAH1,

Assessment of Human Sperm Cells Morphological Parameters

http://dx.doi.org/10.5772/intechopen.71413

61

Figure 7. Spermatozoon with multiple abnormalities. Round head, bent neck and duplicate tail. Bar = 10 μm.

The morphologically abnormal spermatozoon can have multiple defects too (Figure 7). The most irregularly shaped, multi-tailed spermatozoa is associated with severe male infertility. The multiple gene mutations, high rates of polyploidy and aneuploidy and reduced motility of

DNAJB13, CFAP43 and CFAP44) [70–73].

spermatozoa have been described in these cases [48, 49].

Figure 6. The residual cytoplasm on the spermatozoa tail. Bar = 10 μm.

Cytoplasmic residues are large, irregular material along the midpiece and can indicate the abnormal spermiogenesis (Figure 6). They form as outcome of incomplete cytoplasmic extrusion during spermiogenesis in the causes of suppression of FSH and/or androgens, deficiency of cyclin-dependent kinase 16 (CDK16) and organophosphorus pesticide (OP) exposure. In comparison to the cytoplasmic droplet, cytoplasmic residues contain major quantity of cytoplasm enzymes, which produce pathological amounts of reactive oxygen species. These species can negatively affect spermatozoa function including peroxidative damage to the cell membrane, DNA damage, mitochondrial dysfunction, impaired interaction with the female

reproductive tract and lead to male infertility [1, 32, 52, 68, 69].

60 Spermatozoa - Facts and Perspectives

Figure 6. The residual cytoplasm on the spermatozoa tail. Bar = 10 μm.

Figure 7. Spermatozoon with multiple abnormalities. Round head, bent neck and duplicate tail. Bar = 10 μm.

Multiple morphological abnormalities in the same patient's spermatozoa tail structure and ultrastructure which impair motility can show the mutation of some genes (DNAH1, DNAJB13, CFAP43 and CFAP44) [70–73].

The morphologically abnormal spermatozoon can have multiple defects too (Figure 7). The most irregularly shaped, multi-tailed spermatozoa is associated with severe male infertility. The multiple gene mutations, high rates of polyploidy and aneuploidy and reduced motility of spermatozoa have been described in these cases [48, 49].
