Although a lot of efforts have been made for helping the azoospermia patients, there are still 20%-30% unable to obtain sperm suitable for injection. Therefore, repeated punctures were performed to increase the success rate of sperm acquisition in most centers. It was shown that repeated testicular or epididymal puncture has no remarkable effect on the outcomes of pregnancy [6]. However, multiple operations may cause damage to the blood supply of the testicular seminiferous tubules, produce hematoma formation in the testis, and bring about fibrosis of the testicular tissue and even testicular atrophy, which may lead to long-term complications such as male autoimmune response, osteoporosis, insulin resistance and depression [7]. In addition, some azoospermia patients may lose the opportunity to be a biological father in the event of failure of TESA on the day of oocyte collection, and thus have to choose insemination with donor spermatozoa or oocyte cryopreservation, if they did not undergo diagnostic TESA and sperm freezing in advance. Therefore, cryopreservation of testicular sperm is important for fertility preservation in azoospermia patients, the full and effective use of sperm is key to the treatment of obstructive azoospermia. It is commonly believed that freezing sperms after TESA was a safe, economical, and effective way for OA patients [8], and at the same time, the problem of damage mentioned above will be solved. Nevertheless, some scholars were still worried about the pregnancy outcomes of ICSI with frozen testicular sperm and recommended the use of fresh oocytes with fresh testicular sperm as the first-choice treatment [9–10]. Hauser et al.[10] proposed that the use of fresh testicular sperm results in a higher fertilization rate and clinical pregnancy rate than frozen testicular sperm. Therefore, the author insisted that fresh testicular sperm should be considered first for ICSI in patients with virtual azoospermia or cryptozoospermia because of their superior fertility. In 2015, a retrospective study including 110 cycles of TESE-ICSI were conducted by Park et al [11], who confirmed that statistically significant differences were observed in the pregnancy and implantation rates between fresh and frozen testicular spermatozoa group, in spite of similar laboratory outcomes, clinical pregnancy and delivery.
But analogous with the results of our study, most studies have suggested a similar results in terms of fertilization rate, cleavage rate and clinical pregnancy rate in ICSI with frozen testicular sperm [12–14]. OU L et al. [12] found that frozen testicular sperm or fresh testicular sperm extracted from patients with OA have the same 2PN fertilization and pregnancy potential (74.41% vs 76.43% and 46.81% vs 53.39%, respectively). Another retrospective study in 2018 suggested that freezing low-count sperm collected by TESA with a cryoprotectant was an efficient method. The clinical pregnancy rates of ICSI with fresh testicular sperm and frozen testicular sperm were 61.7% and 55.1%, although the pregnancy rate in the fresh sperm group was higher, the difference was not significant. It shows that the two types of sperm are equally reliable for ICSI [15] .
In our study, in addition to focusing on the results of laboratory outcomes, clinical pregnancy and delivery, we further analysed the gender and birth weight of the newborns. There was a resemblance result of sex ratio and average newborn birth weight in the two groups, but in spite of this, the incidence of low-weight newborns significantly increased when frozen testicular sperm was used for ICSI (20.91% vs 8.49%). The average birth weight of low-birth-weight newborns in the frozen and fresh sperm group was 1947g and 2106g respectively. Among them, 17 in the frozen sperm group were twin newborns, and 8 in the other group. To be more specific, in the frozen sperm group, two of the newborns were of very low birth weight (VLBW), and two infants have extremely low birth weight (ELBW)(Table 4). From what has been discussed above, we may reasonably arrive at the conclusion that the use of frozen sperm for ICSI may increase the incidence of low birth weight infants. LBW is one of the leading causes of neonatal and infant morbidity and mortality[16], which is also associated with long-term consequences including cerebral palsy, sensory deficits, cardiovascular and respiratory illnesses. One possible explanation is that resuscitation of frozen sperm has a negative effect on total count and motile count, and poor quality of semen may be related with low birth weight in the offspring [17]. This may also be due to the frozen damage of the sperm during the process of cooling, thawing and cryoprotectant exposure, which might result in changes in the function and structure of sperm [18], bringing about a higher rate of deformed sperm. Higher sperm deformity rate may be contributed to more chances of abnormal expression of imprinting genes, which will cause inefficient placental nutrient supply, resulting in reduction in neonatal birth weight [19]. The above situation is more prominent in multiple pregnancies. LBW is a known complication of multiple gestation [20]. A recently published research revealed that mothers with multiple pregnancies were at a significantly increased risk of delivering babies with LBW, both in the case of preterm and term birth [21]. In our study, the rate of multiple pregnancies in the frozen sperm group was 5% higher than the fresh group. Although there was no significant differences, this may be one of the reasons for the increased low birth weight rate in the freezing group.
Table 4
Detailed Characteristics of Low-Birth-Weight newborns
Case
|
sperm
|
Female age(years)
|
Male age(years)
|
Single/twins
|
Birth weight(g)
|
Gestational age(weeks)
|
1
|
frozen
|
34
|
34
|
single
|
1150
|
31
|
2
|
frozen
|
31
|
31
|
twins
|
2400
|
37
|
3
|
frozen
|
27
|
27
|
single
|
2200
|
37
|
4
|
frozen
|
32
|
39
|
twins
|
2350
|
37
|
5
|
frozen
|
32
|
36
|
single
|
2300
|
37
|
6
|
frozen
|
40
|
44
|
single
|
1850
|
32
|
7
|
frozen
|
27
|
42
|
twins
|
2400
|
36
|
8
|
frozen
|
28
|
31
|
twins
|
2340
|
36
|
9
|
frozen
|
28
|
31
|
twins
|
2440
|
36
|
10
|
frozen
|
32
|
36
|
twins
|
2300
|
37
|
11
|
frozen
|
31
|
36
|
twins
|
2200
|
39
|
12
|
frozen
|
24
|
32
|
single
|
2400
|
37
|
13
|
frozen
|
31
|
34
|
twins
|
1800
|
32
|
14
|
frozen
|
31
|
34
|
twins
|
1600
|
32
|
15
|
frozen
|
26
|
25
|
twins
|
1700
|
30
|
16
|
frozen
|
26
|
25
|
twins
|
1400
|
30
|
17
|
frozen
|
30
|
34
|
twins
|
700
|
34
|
18
|
frozen
|
30
|
34
|
twins
|
700
|
34
|
19
|
frozen
|
26
|
28
|
twins
|
2350
|
37
|
20
|
frozen
|
37
|
42
|
single
|
2300
|
36
|
21
|
frozen
|
31
|
33
|
twins
|
2490
|
38
|
22
|
frozen
|
24
|
27
|
twins
|
1600
|
31
|
23
|
frozen
|
24
|
27
|
twins
|
1800
|
31
|
24
|
fresh
|
25
|
32
|
twins
|
1750
|
33
|
25
|
fresh
|
30
|
33
|
twins
|
2100
|
36
|
26
|
fresh
|
30
|
33
|
twins
|
2250
|
36
|
27
|
fresh
|
24
|
26
|
twins
|
2250
|
38
|
28
|
fresh
|
24
|
26
|
twins
|
2100
|
38
|
29
|
fresh
|
34
|
35
|
twins
|
1850
|
36
|
30
|
fresh
|
29
|
33
|
twins
|
2450
|
37
|
31
|
fresh
|
29
|
33
|
twins
|
2100
|
37
|
32
|
fresh
|
31
|
34
|
single
|
2100
|
33
|
Note There were 23 cases of Low-Birth-Weight newborns in the frozen sperm group(1–23), and 9 cases in the fresh sperm group (24–32). |
The risk of deformity is another problem for newborns after using testicular sperm. There were no cases of fetal or neonatal malformations found in this study. In a population-based cohort study, in which all Danish children were delivered after testicular sperm aspiration (TESA) and fresh embryo transfer in Denmark from 1995 to 2009, the outcome of newborns, including congenital abnormalities after ICSI treatment using epididymal or testicular sperm, has no difference with that after ICSI/IVF treatment using ejaculate sperm or natural conception [22]. A 10-year study in China focusing on the neonatal outcome of children born after ICSI with epididymal or testicular sperm clearly showed that freezing testicular sperm has no effect on the rate of neonatal deformity [23].