Treatment of childhood constipation: a synthesis of systematic reviews and meta-analyses.

Introduction: Constipation occurs in many children and can become chronic. Many grow out of it but for one third, it continues into adulthood. For most patients, there is no identifiable organic disorder and it is classified as functional constipation.Areas covered: In 2016, treatment of childhood constipation was extensively reviewed by Rome IV. This review covers meta-analyses and evidence for treatment of paediatric constipation since 2016 and new emerging treatments.Expert opinion: Since 2016, meta-analyses conclude 1) fibre should be included in a normal diet, but further supplementation does not improve constipation; 2) probiotics may increase stool frequency in children, but evidence from larger RCTs is needed; 3) comparing laxatives, polyethylene glycol (PEG) is superior to placebo, lactulose and milk of magnesia, and 4) appendix stomas are effective and should be considered before surgery. Emerging areas of study include food intolerance, electrical stimulation and faecal microbiota transplant. For research, outcome measures need standardising to allow comparison between studies and allow meta-analyses. To assist this, validated GI instruments have been developed by Rome IV and PedsQl.

Postnatal Germ Cell Development in the Cryptorchid Testis: The Key to Explain Why Early Surgery Decreases the Risk of Malignancy.

PURPOSE: Cryptorchidism is a risk factor for testicular malignancy and surgical treatment lowers this risk. This study aimed to investigate the germ cell behavior in prepubertal cryptorchid testes using immunohistochemical markers for germ cell malignancy to understand how early orchiopexy may possibly prevent cancer developing. MATERIALS AND METHODS: Histology sections from 1,521 consecutive testicular biopsies from 1,134 boys aged 1 month to 16.5 years operated for cryptorchidism were incubated with antibodies including antiplacental-like alkaline phosphatase (PLAP), anti-Oct3/4, anti-C-kit, and anti-D2-40. RESULTS: Oct3/4 and D2-40-positive germ cells are found throughout the first 2 years of life, with declining frequency thereafter. After 2 years, they should have disappeared and may indicate neoplasia. PLAP-positive cells were seen in 57 to 82% and C-kit-positive cells in 5 to 21% of cryptorchid testes between 4 and 13 years. Not until puberty did PLAP and C-kit-positive undifferentiated spermatogonial stem cells vanish. Only 0.3% of the present material had obvious prepubertal intratubular germ cell neoplasia (ITGCN) and they all had syndromic cryptorchidism. An additional three boys (0.3%) older than 2 years had weak Oct3/4 expression in undescended testes, but all cases were D2-40 negative. CONCLUSION: Prepubertal ITGCN was rare and mostly seen in syndromic cryptorchidism. In nonsyndromic cryptorchidism PLAP-positive undifferentiated spermatogonial stem cells persisted in a significant proportion of nontreated undescended testes and they will be especially sensitive to long-lasting abnormally high temperature that may be the single most important cause facilitating the accumulation of mutations during cell replication and the development of ITGCN to be prevented by orchiopexy.

Multiple endocrine neoplasia 2B: Differential increase in enteric nerve subgroups in muscle and mucosa.

Multiple endocrine neoplasia 2B (MEN2B) is a rare syndrome caused by an activating mutation of the RET gene, leading to enteric gangliomatosis. This child presented with constipation at 1-mo old, was diagnosed with MEN2B by rectal biopsy at 4 mo, had thyroidectomy at 9 mo and a colectomy at 4 years. We studied the extent of neuronal and nerve fibre proliferation and which classes of enteric nerves are affected by examining the colon with multiple neuronal antibodies. Resected transverse colon was fixed, frozen, sectioned and processed for fluorescence immunohistochemistry labelling with antibodies against TUJ1, Hu, ChAT, NOS, VIP, SP and CGRP and cKit. Control transverse colon was from the normal margin of Hirschsprung (HSCR) colon (4-year-old) and a child with familial adenomatous polyposis (FAP, 12 year). Myenteric ganglia were increased in size to as wide as the circular muscle. There was a large increase in nerve cells and nerve fibres. ChAT-, NOS-, VIP- and SP-immunoreactive nerve fibres all increased in the myenteric ganglia. NOS-IR nerves preferentially increased in the muscle, while VIP and SP increased in submucosal ganglia and mucosal nerve fibres. The density of ICC was normal. RET overactivation in MEN2B lead to a large increase in intrinsic nerve fibres in the myenteric and submucosal ganglia, with a relative increase in NOS-IR nerve fibres in the circular muscle and VIP and SP in the submucosal ganglia and mucosa. The changes were associated with severe constipation resulting in colectomy at 4 years.

Nuclear transit study in children with chronic faecal soiling after Hirschsprung disease (HSCR) surgery has revealed a group with rapid proximal colonic treatment and possible adverse reactions to food.

BACKGROUND/PURPOSE: Long-term problems with faecal incontinence occur in up to 50 % of patients after pull-through for Hirschsprung disease (HSCR). The cause often remains unknown, leading to empirical treatments. Using nuclear transit study, we found some patients surprisingly had rapid proximal colonic transit, suspicious of occult diarrhoea. We aimed to assess whether these patients had unrecognized adverse reactions to food. METHODS: Patients (n = 10, all males, 9.6 year; 4.25-15.5 years) with persistent faecal incontinence following pull-through for HSCR referred to the senior author and after exclusion of anatomical defects, underwent nuclear transit studies. Most (8) subsequently underwent breath hydrogen tests for sugar malabsorption and were tested for adverse reactions to food. Exclusion diets for protein allergens, lactose or fructose were then trialed. RESULTS: Of the 10 patients with rapid intestinal transit proven on nuclear transit study, breath hydrogen tests for fructose and/or lactose malabsorption were done in 8, and were positive in 7/8 patients. Exclusion diets contributed to either resolution or improvement in faecal incontinence in 9/10 patients. CONCLUSIONS: Rapid transit in the proximal, ganglionated colon may be present in children with faecal incontinence following pull-through for HSCR, possibly secondary to adverse reactions to food. This study suggests that children with post-operative soiling may benefit from a transit study and hydrogen breath tests to diagnose adverse reactions to food caused by sugar malabsorption.

Neurotrophin signaling in a genitofemoral nerve target organ during testicular descent in mice.

BACKGROUND/AIM: It has been proposed that androgens control inguinoscrotal testicular descent via release of calcitonin gene-related peptide (CGRP) from a masculinised genitofemoral nerve (GFN). As there are androgen receptors in the inguinoscrotal fat pad (IFP) during the window of androgen sensitivity (E14-17 in mouse embryos), we tested the hypothesis that neurotrophins in the IFP may masculinise the sensory fibers of the GFN supplying the gubernaculum and IFP prior to gubernacular migration. METHODS: Androgen-receptor knockout (ARKO) and wild-type (WT) mouse embryos were collected at E17, with ethical approval (AEC 734). Sagittal sections of IFP, mammary area and bulbocavernosus (BC) muscle were processed for standard histology and fluorescent immunohistochemistry for ciliary neurotrophic factor (CNTF), ciliary neurotrophic factor receptor (CNTFR) and cell nuclei (DAPI). RESULTS: In the ARKO mouse CNTFR immunoreactivity (CNTFR-IR) was increased in the IFP but decreased in BC. Perinuclear staining of CNTF-IR was seen in mouse sciatic nerve but only weakly in IFP. In the mammary area, also supplied by GFN, there were no differences in IR staining. CONCLUSION: This study found CNTFR-IR in the IFP was negatively regulated by androgen, suggesting that CNTF signaling may be suppressed in GFN sensory nerves to enable CGRP expression for regulating gubernacular migration in the male, but not the female. The indirect action of androgen via the GFN required for testicular descent may be one of the sites of anomalies in the putative multifactorial cause of cryptorchidism.

Oct4-GFP expression during transformation of gonocytes into spermatogonial stem cells in the perinatal mouse testis.

BACKGROUND/AIM: In cryptorchidism perinatal failure to switch off Oct4, a germ cell (GC) marker, may lead to carcinoma in situ. We aimed to analyze Oct4 expression during mouse gonocyte transformation into spermatogonial stem cells (SSC). MATERIALS AND METHODS: Testes from OG2 (Oct4-promoter driven eGFP) mice at embryonic day (E) 17 and postnatal day P0-10 underwent immunohistochemistry and immunoblotting. Antibodies against MVH, AMH, Ki67, and c-Kit were visualized by confocal microscopy. Numbers of Oct4-GFP(+) GC and Oct4-GFP(-) GC/tubule were counted using ImageJ. Data were analyzed using nonparametric one-way ANOVA. RESULTS: GC from E17-P4 were Oct4-GFP(+). Numbers of Oct4-GFP(-) GC/tubule increased from P6-10, whereas Oct4-GFP(+) GC/tubule numbers remained similar between P6 and P10. Sertoli cells proliferated from E17-P10, whereas GC only proliferated from P2. Gonocytes (Oct4-GFP(+)/c-Kit(-)) central in tubules migrated to the basement membrane to become prospermatogonia (Oct4-GFP(+)/c-Kit(-)) and then SSC (Oct4-GFP(+)/c-Kit(+)) from day 4 and further developed into Oct4-GFP(-)/c-Kit(+) at P6. CONCLUSION: In Oct4-GFP mice both centrally located gonocytes and prospermatogonia located at the tubular basement membrane were Oct4-GFP(+)/c-Kit(-) before further developing into SSC (Oct4-GFP(+)/c-Kit(+)). This indicates that Oct4 is important in gonocyte transformation into SSC. Understanding this process will aid GC tumor diagnostics and fertility potential in boys with UDT undergoing orchidopexy.

Disimpaction of children with severe constipation in 3-4 days in a suburban clinic using polyethylene glycol with electrolytes and sodium picosulphate.

AIM: Constipation is a common cause of admission to hospital for disimpaction, as oral laxatives are often inadequate. High-dose oral laxative protocols are used for complete bowel clearance prior to colonoscopy, but have not been reported for treating faecal impaction. The aim of this study was to assess the effectiveness of a high-dose oral protocol using polyethylene glycol with electrolytes (PEG + E) (Movicol Rx) combined with sodium picosulphate (SP) (Dulcolax SP Rx) in faecal impaction in children presenting to a suburban clinic. METHODS: Forty-four children presented with acute/chronic faecal impaction were given six to eight sachets of PEG + E were given on day 1, with decreasing doses on subsequent 3 days, while 15-20 SP drops were given on days 2 and 3. Compliance with medication was achieved using a simple method of motivation, with the child drinking the laxatives in a race. On day 4, PEG + E was reduced to one sachet and SP to 10 drops as an ongoing maintenance dose. Defecation, soiling, diet and water intake was monitored daily for 7 days in a diary. RESULTS: Forty-four children (aged 2-17 years) seen over 8 months were reviewed retrospectively. Children began defecating within 10-12 h reaching a maximum volume of stool/day (four cups) on day 2. All patients were disimpacted successfully and in the week following disimpaction there was no reported faecal soiling or complications. CONCLUSIONS: A high-dose oral protocol combining PEG + E sachets and SP drops successfully and safely disimpacted a cohort of children with acute/chronic constipation presenting to a suburban continence clinic. This protocol appears to be useful to control faecal disimpaction in an outpatient setting, thereby avoiding hospital admission.

Transabdominal electrical stimulation (TES) for the treatment of slow-transit constipation (STC).

Slow-transit constipation (STC) is a newly described subtype of intractable constipation in children which we originally identified with deficiency of substance P in axons supplying the proximal colonic muscle. When nuclear transit studies became available, the patients were found to have slow proximal colonic transit, and responded to antegrade enemas. Using the appendicostomy, we found that there was reduced frequency in propagating sequences throughout the colon. We began testing whether transcutaneous electrical stimulation (TES) could improve motility and symptoms, and over several trials have now shown that TES is remarkably effective in treating children with STC, with long-lasting effects. TES holds promise for treating STC, as well as a range of gastrointestinal motility disorders.

Regulation of testicular descent.

Testicular descent occurs in two morphologically distinct phases, each under different hormonal control from the testis itself. The first phase occurs between 8 and 15 weeks when insulin-like hormone 3 (Insl3) from the Leydig cells stimulates the gubernaculum to swell, thereby anchoring the testis near the future inguinal canal as the foetus grows. Testosterone causes regression of the cranial suspensory ligament to augment the transabdominal phase. The second, or inguinoscrotal phase, occurs between 25 and 35 weeks, when the gubernaculum bulges out of the external ring and migrates to the scrotum, all under control of testosterone. However, androgen acts mostly indirectly via the genitofemoral nerve (GFN), which produces calcitonin gene-related peptide (CGRP) to control the direction of migration. In animal models the androgen receptors are in the inguinoscrotal fat pad, which probably produces a neurotrophin to masculinise the GFN sensory fibres that regulate gubernacular migration. There is little direct evidence that this same process occurs in humans, but CGRP can regulate closure of the processus vaginalis in inguinal hernia, confirming that the GFN probably mediates human testicular descent by a similar mechanism as seen in rodent models. Despite increased understanding about normal testicular descent, the common causes of cryptorchidism remain elusive.

Postnatal germ cell development during mini-puberty in the mouse does not require androgen receptor: implications for managing cryptorchidism.

PURPOSE: Undescended testis leads to infertility and malignancy resulting from aberrant germ cell development. Androgens are proposed to control early germ cell development during the transient postnatal surge of gonadotropins and androgen, known as mini-puberty. We assessed the effect of androgen receptor on perinatal germ cell development in mice. MATERIALS AND METHODS: Testes from androgen receptor knockout mice and wild-type littermates (3 to 4 per group) were collected at embryonic day 17 and postnatal days 0 (birth), 2, 4, 6, 8 and 10 for immunohistochemical analysis. Antibodies against mouse VASA homologue (germ cell marker), antimüllerian hormone (Sertoli cell marker), Ki67 (proliferating cell marker) and DAPI (nuclei) were used and visualized by confocal microscopy. Number of germ cells per tubule, germ cells on the tubular basement membrane and Sertoli cells per tubule, and percentage of proliferating germ cells (Ki67(+)) per tubule and germ cells (Ki67(+)) on the basement membrane on confocal images were counted using Image J, version 1.44 (http://imagej.nih.gov/ij/). Data were analyzed using nonparametric one-way ANOVA with GraphPad Prism® 5.02 software. RESULTS: In wild-type and androgen receptor knockout testes germ cells per tubule decreased from embryonic day 17 to postnatal day 2, then increased normally. Number of mouse VASA homologue positive germ cells per tubule and germ cells on the basement membrane were similar in androgen receptor knockout and wild-type testes (p > 0.05) at each age, and percentages of proliferating germ cells (Ki67(+)) per tubule and proliferating germ cells on the basement membrane were similar at each age (p > 0.05). CONCLUSIONS: Androgen receptors are not required for gonocyte migration from the center of the testicular tubules to the basement membrane and transformation into spermatogonia stem cells up to day 10 in androgen receptor knockout mice. Identifying nonandrogenic factors might improve the fertility potential of boys with undescended testis who are undergoing orchiopexy.

Molecular signals governing cremaster muscle development: clues for cryptorchidism.

BACKGROUND/AIM: Cryptorchidism affects 2-4% of newborn boys. Testicular descent requires the gubernaculum to differentiate into cremaster muscle (CM) during androgen-mediated inguino-scrotal descent, but the cellular mechanisms regulating this remodeling remain elusive. ß-Catenin, a marker of canonical Wnt signaling, promotes myogenic genes and cellular adhesion. We aimed to determine if androgen receptor (AR) blockade altered ß-catenin and its downstream myogenic proteins within the CM. METHOD: Gubernacula from male rats (n=12) and rats treated with anti-androgen, flutamide (n=12) at E19, D0, D2 were processed for immunohistochemistry. Antibodies against ß-catenin, embryonic myosin, and myogenin were visualized by confocal microscopy. RESULTS: At E19, ß-catenin immuno-reactivity (IR) localized to the CM membrane. By D2, cytoplasmic ß-catenin-IR was noted with overall ß-catenin-IR decreasing. Myogenic proteins resided primarily in cells containing ß-catenin on their plasma membrane. Embryonic myosin-IR was high at E19 and then decreased by D2, while myogenin-IR increased. AR blockade increased cytoplasmic ß-catenin at D2 and reduced levels of both myogenic proteins. CONCLUSION: Myogenic proteins are present in CM cells containing ß-catenin. AR blockade did not alter cellular adhesion via ß-catenin. In contrast, blocking AR prevented ß-catenin entering the nucleus and impaired CM myogenesis. Mutations in this pathway may result in idiopathic cryptorchidism.

Immunofluorescent analysis of testicular biopsies with germ cell and Sertoli cell markers shows significant MVH negative germ cell depletion with older age at orchiopexy.

PURPOSE: Undescended testis is the most common defect in male newborns. This condition is associated with increased risks of infertility and testicular malignancy due to abnormal germ cell development in the testes. Early surgery may limit such risks. We analyzed germ cell development vs age at orchiopexy using a germ cell marker and a Sertoli cell marker on testicular biopsies. MATERIALS AND METHODS: A total of 22 testicular biopsies at orchiopexy in 20 patients 5 to 24.5 months old were fixed and embedded in paraffin. Sections were processed and labeled with AMH antibody for Sertoli cells and MVH antibody for germ cells for immunofluorescent histochemical analysis. Confocal images were counted using ImageJ (National Institutes of Health, Bethesda, Maryland) for germ cells and testicular tubules. The data were analyzed using linear regression. RESULTS: Sertoli cells were clearly distinguished from MVH positive and negative germ cells located centrally or on basement membranes of tubules. Percentage of tubules with MVH negative germ cells significantly decreased with increasing age at orchiopexy (ß = -0.03, p = 0.03). Total tubular numbers and "empty" tubules without germ cells significantly increased with age at orchiopexy (ß = 1.15, p = 0.02 and ß = 0.44, p = 0.04, respectively). CONCLUSIONS: AMH antibody distinguished Sertoli cells from germ cells, and MVH antibody distinguished 2 types of germ cells at different developmental stages. Biopsy at orchiopexy in older patients showed significant germ cell depletion. These results lend support to early surgery to optimize germ cell number.

Hoxa-11 maintains cell proliferation in the mouse gubernaculum to facilitate testicular descent.

INTRODUCTION: The gubernaculum is a structure vital for guiding testicular descent. The Homeobox gene, Hoxa-11, is involved in patterning embryonic structures and is necessary for gubernacular development, as Hoxa-11 knock-out mice exhibit abnormal gubernacula and undescended testes. We aimed to elucidate how testicular descent fails by examining cell proliferation and androgen receptor (AR) expression in Hoxa-11 KO mice gubernacula. METHODS: Postnatal day 2 wild type (n=6) and Hoxa-11 KO mice (n=6), were prepared for immunohistochemistry and confocal microscopy using antibodies against androgen receptor, slow skeletal myosin (My32), and Ki67, a marker of cell proliferation. RESULTS: The gubernacula of Hoxa-11 KO mice were hypocellular compared with WT. AR was present in the gubernaculum and abutting inguinal fat pad in both WT and Hoxa-11 KO with no difference in expression. Slow skeletal myosin was present in a clear 'swirl' in the growth centre of WT animals which was absent in the Hoxa-11 KO mice. Ki67, expressed in the growth centre and cremaster muscle in WT, was greatly decreased in Hoxa-11 KO. CONCLUSION: Hoxa-11 may regulate fibroblast proliferation in the gubernaculum, as it does in human uterosacral ligaments, allowing formation of the 'growth centre' within the bulb and facilitating myogenesis and elongation to the scrotum. Polymorphisms in Hoxa-11 may contribute to the aetiology of human cryptorchidism.

Wnt signalling in testicular descent: a candidate mechanism for cryptorchidism in Robinow syndrome.

BACKGROUND/AIMS: Robinow syndrome is caused by mutations in Wnt-5a or its receptor Ror2 and can lead to cryptorchidism, though the mechanisms are unclear. Wnt-5a knock-out mice fail to undergo gubernacular swelling, similar to insulin-like hormone 3 (INSl-3) knock-out mice. We aimed to characterise Wnt-5a and Ror2 expression in rat gubernacula to better understand how Wnt-5a signalling affects testicular descent. METHODS: Sprague-Dawley rats (n = 27) were collected with ethics approval (A644) at embryonic days (E) 15, 17, 19 and postnatal day (D) 2. Control and antiandrogen-treated groups were processed for immunohistochemistry for Wnt-5a, Ror2 and ß-catenin. Sagittal sections were examined using confocal microscopy. RESULTS: Wnt-5a and Ror2 were strongly expressed in the gubernacular bulb at E17 controls, their levels declining at E19 and almost absent by D2. Wnt-5a significantly co-localised with the important transcription factor ß-catenin at E17. There was no obvious difference in staining with androgen blockade. CONCLUSION: Wnt-5a, through Ror2 and ß-catenin may play a vital role in regulating the gubernacular swelling reaction downstream of INSL-3. Human mutations in Wnt-5a or Ror2 could prevent early gubernacular growth, as suggested by undescended testes in 70% of patients with Robinow Syndrome.

The regulation of testicular descent and the effects of cryptorchidism.

The first half of this review examines the boundary between endocrinology and embryonic development, with the aim of highlighting the way hormones and signaling systems regulate the complex morphological changes to enable the intra-abdominal fetal testes to reach the scrotum. The genitoinguinal ligament, or gubernaculum, first enlarges to hold the testis near the groin, and then it develops limb-bud-like properties and migrates across the pubic region to reach the scrotum. Recent advances show key roles for insulin-like hormone 3 in the first step, with androgen and the genitofemoral nerve involved in the second step. The mammary line may also be involved in initiating the migration. The key events in early postnatal germ cell development are then reviewed because there is mounting evidence for this to be crucial in preventing infertility and malignancy later in life. We review the recent advances in what is known about the etiology of cryptorchidism and summarize the syndromes where a specific molecular cause has been found. Finally, we cover the recent literature on timing of surgery, the issues around acquired cryptorchidism, and the limited role of hormone therapy. We conclude with some observations about the differences between animal models and baby boys with cryptorchidism.

Transabdominal electrical stimulation increases colonic propagating pressure waves in paediatric slow transit constipation.

BACKGROUND AND AIMS: In slow-transit constipation (STC) pancolonic manometry shows significantly reduced antegrade propagating sequences (PS) and no response to physiological stimuli. This study aimed to determine whether transcutaneous electrical stimulation using interferential current (IFC) applied to the abdomen increased colonic PS in STC children. METHODS: Eight children (8-18 years) with confirmed STC had 24-h colonic manometry using a water-perfused, 8-channel catheter with 7.5 cm sidehole distance introduced via appendix stomas. They then received 12 sessions (20 min/3× per week) of IFC stimulation (2 paraspinal and 2 abdominal electrodes), applied at a comfortable intensity (<40 mA, carrier frequency 4 kHz, varying beat frequency 80-150 Hz). Colonic manometry was repeated 2 (n=6) and 7 (n=2) months after IFC. RESULTS: IFC significantly increased frequency of total PS/24h (mean ± SEM, pre 78 ± 34 vs post 210 ± 62, p=0.008, n=7), antegrade PS/24h (43 ± 16 vs 112 ± 20, p=0.01) and high amplitude PS (HAPS/24h, 5 ± 2:10 ± 3, p=0.04), with amplitude, velocity, or propagating distance unchanged. There was increased activity on waking and 4/8 ceased using antegrade continence enemas. CONCLUSIONS AND INFERENCES: Transcutaneous IFC increased colonic PS frequency in STC children with effects lasting 2-7 months. IFC may provide a treatment for children with treatment-resistant STC.

Is matrix metalloproteinase required in postnatal testicular tubules for germ cell maturation?

BACKGROUND/AIM: Cryptorchidism may cause infertility by failed transformation of neonatal gonocytes into adult dark spermatogonia, the putative stem cells for spermatogenesis. Gonocytes migrate centrifugally to the tubular basement membrane to become adult dark spermatogonia. Regulation of this transformation remains unknown. We aimed to investigate neonatal rodent testis matrix metalloproteinase (MMP) production to see whether MMPs loosen extracellular matrix between Sertoli cells to facilitate gonocyte movement. METHODS: Sprague-Dawley rat testes (n = 4-6 per group) were collected at embryonic day 19 (E19) and postnatal (P) days P0 to 10 for immunohistochemistry. Immunofluorescent confocal images were captured for presence of membrane type 1 MMP (MT1-MMP), matrix metalloproteinase 2 (MMP2), tissue inhibitor of metalloproteinase 2 (TIMP2), mouse VASA homologue, anti-Müllerian hormone, and androgen receptor in tissue sections. Testicular proteins were analyzed by immunoblotting. RESULTS: Membrane type 1 MMP was strongly present in gonocytes at E19 then decreased, whereas it increased in testicular somatic cells from P0 to P10. Testicular protein levels of MT1-MMP, MMP2, and androgen receptor were constant from E19 to P10. Anti-Müllerian hormone protein sharply decreased after P2, whereas TIMP2 gradually increased from E19 to P10. Gonocytes migrated to basement membrane at P2 to P6. CONCLUSION: Membrane type 1 MMP, MMP2, and TIMP2 were present in testis from E19 to P10 during gonocyte migration and transformation into spermatogenic stem cells. Increased knowledge about germ cell development may aid efforts to improve fertility in cryptorchidism.