Gene: 268Name: Twg-pendingFamily: Growth FactorSubfamily: TwistedAccession: AF292033GI: 9837569

Gene 268
Summary of Phenotypic Analysis


Changes related to genotype:

ES cells derived from the 129/SvJ x 129/Sv-CP mouse substrain were used to generate chimeric mice. F1 mice were generated by breeding with C57BL/6 females. F2 homozygous mutant mice were produced by intercrossing F1 heterozygous males and females.

Wild-type control mice, as well as homozygous mutant and heterozygous mutant mice, were evaluated by the following examinations or tests:

When compared to age- and gender-matched wild-type control mice, homozygous mutant mice frequently had kinky tails at 49 days of age, and infrequently at 300 days of age.  Body length was short and body weight was low in homozygous mutant males at 300 days of age.  Aging metrics showed trends for short body length, low body weight, and low body weight / body length ratio in homozygous mutant males and, to a lesser degree, in homozygous mutant females at 49, 90, 180, and 300 days of age.

Gene 268
Behavior

Changes related to genotype:

There were no significant differences detected in the homozygous mutant animals when compared with age- and gender-matched wild-type control mice.

Homozygous mutant and wild-type control mice were evaluated for phenotypic changes by testing on six behavioral tasks: Open field test, Tail suspension test, Rotarod test, Hot plate test, Startle/PPI, and Metrazol test.

Mouse ID numbers are as follows:

10 homozygous mutant males (68299, 51253, 68298, 59521, 68295, 51314, 68305, 68307, 68296, 59725)
10 wild-type control males 
(68300, 59518, 68311, 68297, 71660, 65748, 59723, 59724, 68303, 71657)

ES cells derived from the 129/SvJ x 129/Sv-CP mouse substrain were used to generate chimeric mice.  F1 mice were generated by breeding with C57BL/6 females. The resultant F1N0 heterozygotes were backcrossed to C57BL/6 mice to generate F1N1 heterozygotes.  F2N1 homozygous mutant mice were produced by intercrossing F1N1 heterozygous males and females.

Behavior Findings:
There were no genotype-related or biologically significant differences noted between homozygous mutant and wild-type control mice for any of the parameters evaluated during behavior testing.

Gene 268
Fertility


Both homozygous males and females were fertile. Their progeny were viable until weaning, which suggests no abnormalities in the ability of the mutant females to nurture their pups.

Two to three homozygous mutant mice of each gender were set up in a fertility mating with a wild-type mate at seven to eight weeks of age. The numbers of pups born from one to three litters were recorded. Three weeks later, the live pups were counted and weaned.

Mouse ID numbers are as follows:

3 homozygous mutant males (10827, 28909, 50962)
2 homozygous mutant females (9138, 28906)

Gene 268
Expression Summary

RT-PCR Summary:
RNA transcripts are detectable in all tissues analyzed: brain, cortex, subcortical region, cerebellum, brainstem, olfactory bulb, eye, heart, lung, liver, pancreas, kidneys, spleen, thymus, lymph nodes, bone marrow, skin, gallbladder, urinary bladder, pituitary gland, adrenal gland, salivary gland, skeletal muscle, tongue, stomach, small intestine, large intestine, cecum, testis, epididymis, seminal vesicle, coagulating gland, prostate gland, ovary and uterus.

Gene 268
Histopathology

There were no significant differences detected in the homozygous mutant and heterozygous mutant animals when compared with age- and gender-matched wild-type control mice.

The following mice were evaluated by histopathological examination.

49 Day Cohort Mouse ID numbers are as follows:
3 homozygous mutant females (10796, 10799, 10802)
3 homozygous mutant males (10828, 10829, 10834)
1 heterozygous mutant female (10801)
1 heterozygous mutant male (10832)
1 wild-type control female (10798)
1 wild-type control male (10833)

300 Day Cohort Mouse ID numbers are as follows:
4 homozygous mutant females (10699, 10702, 11656, 13263)
5 homozygous mutant males (10827, 11655, 11657, 11658, 15612)
3 wild-type control females (10701, 13264, 51411)
3 wild-type control males (11212, 14152, 15084)

No Significant Abnormalities:
Tissues examined and considered to have no genotypically significant abnormality: brain, pituitary gland, ears, nasal cavity, salivary glands, oral cavity, lymph nodes, aorta, lungs, gallbladder, pancreas, spleen, kidneys, urinary bladder, stomach, small and large intestines, larynx, esophagus, trachea, thyroid gland, thymus gland, tongue, skeletal muscle, sciatic nerve, mammary glands, vertebrae, spinal cord, bone (skull, sternum, femur, tibia, and stifle joint), reproductive tract (including gonads), eyes, Harderian glands, integumentary system (skin and either clitoral or preputial glands), and bone marrow.

Bone marrow was evaluated in sections of sternum, vertebrae, and/or femur and tibia. Marrow cellularity, numbers of megakaryocytes, and myeloid:erythroid (M:E) ratio are within normal limits. Myeloid and erythroid precursors show a normal maturation sequence.

Incidental lesions were present in some tissues, including Harderian gland adenomas in 2 homozygous mutant mice (female 13263, male 15612) and a cystic bulbourethral gland in a homozygous mutant male (10827). These findings are considered to represent background lesions occasionally seen in this strain of mice, lesions due to spontaneous disease, age-related lesions, lesions due to procedural artifacts, and/or lesions of a nonspecific etiology. They are not considered to be genotype related.

Gene 268
Necropsy 

Changes related to genotype:

The following mice were necropsied. Body weight, body length, and organ weights were obtained, and gross pathological findings were recorded.

49 Day Cohort Mouse ID numbers are as follows:
5 homozygous mutant females (10799, 10802, 16040, 16042, 17646)
6 homozygous mutant males (10828, 10829, 10834, 18210, 19478,19479)
2 heterozygous mutant females (10801, 16041) 
2 heterozygous mutant males (10832, 18208) 
3 wild-type control females (10798, 17458, 17459)
3 wild-type control males (10833, 18211, 19425)

300 Day Cohort Mouse ID numbers are as follows:
4 homozygous mutant females (10699, 10702, 11656, 13263)
5 homozygous mutant males (10827, 11655, 11657, 11658, 15612)
3 wild-type control females (10701, 13264, 51411)
3 wild-type control males (11212, 14152, 15084)

Mice were examined for the following observables: adrenal glands, body length, body weight, bone marrow, bone - cranium, bone - femur, bone - sternum, bone - stifle joint, bone - vertebral column, brain, cecum, colon, duodenum, epididymis - seminal vesicle, esophagus, eyes, gallbladder, general appearance, Harderian glands, heart, heart weight, ileum, jejunum, kidney weight, kidneys, liver, liver weight, lungs, lymph nodes, mesentery, ovaries, pancreas, penis, salivary glands, sciatic nerve, scrotum, skeletal muscle, skin, skinned mouse, spleen, spleen weight, stomach, testes, testes - epididymis weight, thymus, thymus weight, tongue, trachea, urinary bladder, urine, uterus and vagina. (Gender-specific observables apply to the appropriate gender.)

Necropsy Findings:

There were no genotype-related or biologically significant differences noted between mutant and wild-type control mice for any of the parameters evaluated at necropsy.  Incidental lesions were present in some tissues, including a mass from the inguinal region in a homozygous mutant male (10827) that was a cystic bulbourethral gland histologically.  These findings were considered to represent background lesions occasionally seen in this strain of mice, lesions due to spontaneous disease, age-related lesions, lesions due to procedural artifacts, and/or lesions of a nonspecific etiology.  They were not considered to be related to genotype.

Body and Organ Weight Findings:

When compared to age- and gender-matched wild-type control mice in the 300 day cohort, body length was short and body weight was low for 4 of 5 homozygous mutant males (10827, 11655, 11658, 15612).  These differences were regarded as related to genotype.  Other differences in body length, body weight, organ weights, and/or organ weight to body weight ratios were present between individual mice.  The variability between mice for these differences usually fell within our historical reference ranges and was not correlated with genotype.

Gene 268
Clinical Chemistry

There were no significant differences detected in the homozygous mutant and heterozygous mutant animals when compared with age- and gender-matched wild-type control mice.

Serum samples from the following mice were evaluated by a clinical biochemistry panel. The data were compiled from N0F2 and N1F2 generations.

49 Day Cohort Mouse ID numbers are as follows:
3 homozygous mutant females (16042, 77285, 83340)
3 homozygous mutant males (74894, 75995, 78632)
1 heterozygous mutant female (77655)
1 heterozygous mutant male (78633)
2 wild-type control females (74907, 83338)
2 wild-type control males (74896, 74902)

90 Day Cohort Mouse ID numbers are as follows:
1 homozygous mutant female (14157)
4 homozygous mutant males (14151, 14153, 14155, 46975)
2 wild-type control females (14159, 14934)
1 wild-type control male (14152)

180 Day Cohort Mouse ID numbers are as follows:
4 homozygous mutant females (13263, 14157, 14935, 15078)
4 homozygous mutant males (14151, 14153, 15612, 46975)
4 wild-type control females (13264, 14159, 14934, 15081)
3 wild-type control males (14152, 15082, 15084)

300-360 Day Cohort Mouse ID numbers are as follows:
4 homozygous mutant females (13263, 50952, 50954, 51412)
4 homozygous mutant males (10827, 11655, 11657, 15612)
4 wild-type control females (13264, 51409, 51411, 57775)
3 wild-type control males (11212, 14152, 57781)

Values for the various analytes evaluated were generally similar between homozygous mutant, heterozygous mutant, and wild-type control mice. Variations in clinical chemistry values, if present, were not consistent with genotype and, thus, were not considered phenotypically relevant.

Gene 268
Hematology

There were no significant differences detected in the homozygous mutant and heterozygous mutant animals when compared with age- and gender-matched wild-type control mice.

Blood samples from the following mice were evaluated by a complete blood count and differential cell count. The data were compiled from N0F2 and N1F2 generations.

49 Day Cohort Mouse ID numbers are as follows:
3 homozygous mutant females (74906, 76000, 83340)
3 homozygous mutant males (74894, 75995, 78632)
1 heterozygous mutant female (77655)
1 heterozygous mutant male (78633)
2 wild-type control females (74907, 83338)
2 wild-type control males (74896, 74902)

90 Day Cohort Mouse ID numbers are as follows:
1 homozygous mutant female (46962)

180 Day Cohort Mouse ID numbers are as follows:
2 homozygous mutant females (13263, 14935)
2 homozygous mutant males (14151, 15612)
4 wild-type control females (13264, 14159, 14934, 15081)
2 wild-type control males (14152, 15082)

300-360 Day Cohort Mouse ID numbers are as follows:
4 homozygous mutant females (11656, 13263, 50954, 51408)
4 homozygous mutant males (10827, 11655, 15612, 58101)
4 wild-type control females (13264, 51409, 51410, 51411)
3 wild-type control males (11212, 14152, 15084)

Although minor variations of hematological values were present in some animals, these changes were not consistent with genotype and, thus, were not considered phenotypically relevant.

Gene 268
Physical Examination

Changes related to genotype:

The following mice were evaluated by physical examination.

49 Day Cohort Mouse ID numbers are as follows:
5 homozygous mutant females (10796, 10799, 16040, 16042, 17646)
4 homozygous mutant males (10829, 18210, 19478, 19479)
1 heterozygous mutant female (16041)
1 heterozygous mutant male (18208)
2 wild-type control females (17458, 17459)
2 wild-type control males (18211, 19425)

300 Day Cohort Mouse ID numbers are as follows:
4 homozygous mutant females (10699, 10702, 11656, 13263)
5 homozygous mutant males (10827, 11655, 11657, 11658, 15612)
3 wild-type control females (10701, 13264, 51411)
3 wild-type control males (11212, 14152, 15084)

Mice were examined for the following observables: anus, behavior, body shape, claws, coat - fur, coat color - back, coat color - belly, ear - left, ear - right, eye - left, eye - right, eye color - left, eye color - right, feces, forelimb - left, forelimb - right, forelimb number of amputated digits - left, forelimb number of amputated digits - right, forelimb number of digits - left, forelimb number of digits - right, general appearance, genitals - female, genitals - male, hair type, head shape, hindlimb - left, hindlimb - right, hindlimb number of amputated digits - left, hindlimb number of amputated digits - right, hindlimb number of digits - left, hindlimb number of digits - right, injuries, lesions, limb shape, locomotor, lumps - masses, mammary glands, mice in cage, respiration, skin appearance, snout, swelling - joints, tail, teeth color, teeth length, urine, and whiskers. (Gender-specific observables apply to appropriate gender.)

At 49 days, when compared to age- and gender-matched wild-type control mice, 5 of the 9 homozygous mice examined, 2 females (10796, 10799) and 3 males (10829, 19478, 19479), had kinky tails.

There were no other genotype-related or biologically significant differences noted between mutant and wild-type control mice at 49 days for any of the parameters evaluated at physcial examination.

At 300 days (actual ages 303 to 450 days), when compared to age- and gender-matched wild-type control mice, 1 male (10827) of the 9 homozygous mutant mice had a kinky tail.

Occasional other differences in physical features of individual mutant mice were present when compared to wild-type control mice. A mass in one homozygous mutant male (10827) was a bulbourethral gland cyst histologically. Three homozygous mutant mice (10827, 11655, 11656) had exophthalmos, which was associated with Harderian gland inflammation in one female (11656), but was not associated with histologic lesions in the other 2 mice. One homozygous mutant male (15612) was squinting, and this was associated with a Harderian gland adenoma. These findings are considered to represent individual variability, background features occasionally seen in this strain of mice, findings due to spontaneous disease, age-related findings, procedural artifacts, and/or findings of a nonspecific etiology. However, none of these differences was regarded as genotype related.

Gene 268
Aging Metrics

Changes related to genotype:

Body weights and body lengths were measured for mice at  20, 49, 90, 180 and 300 days of age.

49 Day Cohort Mouse ID numbers are as follows:
8 homozygous mutant females (10699, 10702, 11656, 14157, 14935, 15078, 46962, 47116)
9 homozygous mutant males (11655, 11657, 11658, 14151, 14153, 14155, 15612, 46975, 47109)
8 heterozygous mutant females (10700, 11209, 11210, 11211, 14156, 15077, 15079, 15080)
7 heterozygous mutant males (11213, 11214, 11215, 11216, 14154, 15083, 15085)
4 wild-type control females (10701, 14159, 14934, 15081)
4 wild-type control males (11212, 14152, 15082, 15084)

90 Day Cohort Mouse ID numbers are as follows:
6 homozygous mutant females (10699, 10702, 11656, 13263, 46962, 47116)
5 homozygous mutant males (11655, 11657, 11658, 46975, 47109)
5 heterozygous mutant females (10700, 11209, 11210, 11211, 12873)
4 heterozygous mutant males (11213, 11214, 11215, 11216)
2 wild-type control females (10701, 13264)
1 wild-type control male (11212)

180 Day Cohort Mouse ID numbers are as follows:
6 homozygous mutant females (13263, 14157, 14935, 15078, 46962, 47116)
5 homozygous mutant males (14151, 14153, 15612, 46975, 47109)
4 wild-type control females (13264, 14159, 14934, 15081)
3 wild-type control males (14152, 15082, 15084)

300 Day Cohort Mouse ID numbers are as follows:
7 homozygous mutant females (10699, 10702, 11656, 13263, 14157, 14935, 15078)
7 homozygous mutant males (10827, 11655, 11657, 11658, 14151, 14153, 15612)
6 wild-type control females (10701, 13264, 14159, 14934, 15081, 51411)
4 wild-type control males (11212, 14152, 15082, 15084)

Body Weight and Length Findings:
When compared to age- (49, 90, 180, and 300 days) and gender-matched wild-type control mice, the homzygous mutant mice had lower values for all 3 parameters (body weight, body length, and body weight/body length ratio) at all time points. Generally the differences between homozygous mutant and matched wild-type control mice were greater for the males than for the females. The body weight and the body weight/body length ratio decreased for homozygous mutant males after 180 days of age. For each of the 3 parameters, the magnitude of the difference between wild-type control and homozygous mutant males was greatest at 300 days of age.