- Email: [email protected]
myo-inositol content of the lens in hereditary cataract mice
Exy. Eye Res. 26, (1978) 119-122
LETTER
TO THE
EDITORS
myo-Inositol Content of the Lens in Hereditary Cataract Mice The free myo-inositol content in the mammalian eye lens is rather high and varies with pathologic states. van Heyningen (1957) reported the decreased mvo-inositol level in the completely opaque lens of the rabbit after X-ray irradiation. Low mvoinositol content, was also observed in lenses of sugar. diabetic and senile cataracts. Studies on the mechanism of developmental and morphological changes in hereditar? cataract have Ijeen performed using the Nakano mouse (Nakano et al., 1960; Iwata and Kinoshita. 1971; Hamai, Fukui and Kuwabara, 1974; Fukui. Gbazawa aml Kinoshita, 1976). We reported here changes of free and lmuntl myo-inositol contmts in lenses of the Nakano cataract mice. _A uralr Nakano mouse was crossed with a BALB/c female. Their litters were mate(l to obtain hon~ozygous mice which developed cataract within a month after birth. Their oRspring were used. BALB/c mice served as normal controls. myo-Inositol was assayed by gas-chromatography as its trimethylsilyl et’her by the method of Yamakawa and Ucda (1964). Gas-chromatographic determination was carried out with R”,, OV-1 on Gas-Chrom P and 5% Ucon LB-550 X on Gas-C’hrom CLH in a glass colunm 3 mm x 2 m. To determine bound myo-inositol in lipids, dried lenses were extracted with chloroform methanol mixture (2 : 1, v/v) according to Folch. Lee and Sloane (1957). The est’ract was once washed wit*h one fifth volume of water to remove free
Age (days) FIG. 1. Concentration of free myo-inositol in the lens of normal (o-0) Each value represents an average of two or three determinations pooled
and cataract (c-9) lenses.
mice.
I20
1.:. \\..\I).\
15T .\I..
sugar atltl polvols. The tlrietl lipid fraction \ws nt~+Jtanolizct1I\-itlt (Fi”,,) iJJl(ltreated with IK AK, tlriwl. ildJICl tritttcth~lsilylatetl. The
COJltf?Jlt
Of
Ill~O-itlOSitO1
it1
th?
hh.
kidney.
tf?StiS
;tll(I
Itl~~t~llil~tl~ll IIC ‘I
hvtht \VilS ;lt il. ~ittll~ilt
level in normal aid cataract mice. In the lens. howt~vcr, thc$ tiiyo-itiositol cotttt~tttN was alqnwximatel,y 5W,, 1on.er in the cataract than in the nortttal tttottw OII ~4; weight I)asis. It was evident t,liat Oh(~hcreditarv tleficiencv afYectot1t,ltt* I(~\YI 01’f’rc~ myo-inositol only in the lens but, not; in ot.lwr-orgarls. I\;-at,a and Kittosltit,;t ( 197I) also oltserved dcficiencv of Nat K ATYaw onlv in the l~~nses of Nakano cataract. IIIIW. Figure l(a) show tler;~lopttrental chat1geh of’ tttvo-inositol _ 1~~~~1 in IJotlt nortttal ;rt~tl cataract, JIJOIJW lenses.The level was the samein I)oth Ictw~sttntjil 10 da!;> :tf%c~r I)irt,tt. Therea,fter. the tnyo-inositol content8 of ttortnal tttonse Irns incrcwecl cotttitluott,sl! to reach ntore than 6 &lenh. In the cataract iJ~01tw lens. howc~vt~l~. i?hc c~lttt~f’tlt increased gratluallv until 30 (tars of age and was then inaittt~aittc~l itt, it htiiitillii;tI value of al)out( I &lrrts. On t’hc: Ilt#h day when Iwth tnicc opettwl t#hc>irt’vt~ atul cataract niicc had transparent8 cbw, thrx lens niyo-inositol l(>wl ~vits sligltt,l;; iowr in the cataract JJ~OIJSCL Ohan in the norrttal. Thus thrl tlly-ill~JShl~ cottttqt of t It,% cataract tttome lens was 60” I, of the JloJ’JJla~ kW?l at f3 \V~xkS (Jf W&y! 1vhCll “pill-tlV;t(l” opacity appeared and only 20” ,, of the normal at 3 months. Whew esprtwe~l on ;I cln weight basis.the myo-inositol content continued to increaseat) it sitttilar rate III) to tttc: age of 50 days in both normal an(l cataract mice with a lag lwriotl of whoub20 cla\-s in the lat,w [Fig. 1(tt)]. Howevcrt it r~w~ainetlat the sattle thrwaftrr in the (~ttt~>trit.ct lens, suggestingthat leveling off to putit]) acbivitv ma\- occur in th(slcrls atj t,ltis agc~. ~OJTeh~i~JJl of rtiyo-inositol level a.11~1 opacity ;,f cataract Icws w-as t’XiJ,ttliJlt?(~ (‘lht)lfs 1). Two litters consisting of tttice with letlses varying itt opacity ant1 OJKS with (.‘l+BitJ’
No. of exp.
1
2
Age (days)
40
34
x0. of lenses
Lens weinht c (w) wet dry
2 4
423 3.1
2
3.2
2 4 4
* Determined macroscopically. - Almost normal, + slight
Degree (right)
2.9
I.21 I.25 1.1” 0.94
++ + i- t-
2.7 24
1.17 0~90
i.+--
cortical
opacity,
of opacity (left)
-t
-k-!-t++
1.7 I.3 14 0.9
-
1.7 9.7
-
-t + marked
+++
opacity
pg of myo-lnositol (/lens) (/rng dry wt.)
and + + + conlplete
140 Ia4
WY!l U.96 143 Cl-SY
opacity.
lenseswere specially selected and divided into four groups. It was remarkablr~ that the lensesfrom these mice, even though transparent, were far lower in u-eight than those of normal mice (5 mg or more) of the sameages. This suggeststhat distur1tattce of lens growth may occur prior to opacification. The clear lens had 1.i pg of ntvc~inositol content corresponding to 70-800,; of the normal. The content was tttarkedly lower in the opaque lenses than in control lenses. It is of interest that the myoinositol content was decreasedwith degree of opacification.
.\IYO-INOSITOL
IX
HEREDITARY
(‘ATARA(“I
121
13otlr lipids and bound myo-inositol content were higher in the cataractous lens thm the normal when expressed as per unit of dry weight (Table II). However, taking account of smaller size and higher water content in the cataract lens. no differelm was found in bound myo-inositol content per lens between cataract anti H~I-I~:L~ mice. It should be pointed out that the amount of hound nlyo-inositol is apparentl,y a small fraction of total mnyo-inositol. In the normd mouse lens free autl Itountl uryo-inositol content was 3.87 and OC ~nml~g wet weight. respectively. In
the cataract lens they were 1.85 and 0.04 +lol, respectively. The ratio tJf IK~LUK~ t,cj myo-inositol was O*52o/o in the normal lens and 2.16” ,, in the cataract lens. Thr Immtl inositol content in the mouse lens was lower by two figures thm that, in rat, tissws reported by Wells, Pittman and Wells (1965). The cataract luoust! lens is sharply contrastStd with the normal one in markedly lowr free inositol cont,cnt. Since phospholipitls are important constituents of t’he nwmhrane syst,em, it is concciva.l)le that t~ht~ membrane system inclutlin g the lens capsule tlicl not, untlrrpo intense conqmitional changes. If there are arl,v defects in the nwnIbmne systcrn, tNlwv we rather functional disorders. frer
KI~FFR~ESCES , , Folch.
J., Let, AI. and Sloane, S. G. H. (195i). -1 simple method for the isolatiorl of total lipids from animal tissues. J. Hid. Chum. 226, 41%319. Hamai, T., .Fukui, H. N. and Kua-abara, ‘I’. (1974). Morphology of hereditary
JSxp. Eye Hes. 18,
at~tl p~lrijicntiorl ~IOIIW
cxt;lrar.t.
537-415.
ran Heyningen, R.. (1957). Mesoinositul in the lens of mammalian eyes. UiocJwu. J. 65, 24 S. Fukui. H. S.. Obazawa, H. and liinoshita. -7. H. (19ili). Lens gron th in the Sakano ~OLISC. Invest. Ophthrrlmol. 15, 422--S. Iweta, S. and Kinoshita. J. H. (19il). !Ucchanism of tlelelopment of hereditary cataract in rrriw. I/west. Ophthdmol. 10, :il)l-1’. Sakano, K., Yamamoto. S.. Kutsukake, C:., Ogawa, H., Sakajima, .I. and Takano. E. (LRtiO). Hereditary cataract in mice. .Jap. J. C’lin. Ophthd. 14, 196-200. Wells, \V. \V., I’ittman, T. A. and IVeIls, H. J. (1965). & uantibtive analysis of myo-inoaitol in rat tissue by gas-liquid chromatography. Annlyt. Hiochem. 10, 450-8. Tamakawa, T. and Veda. S. (1964). C;as-liyuid chromatography of carbohydrates. J~rl/. J. h’.rp.
Jfed. 34, 3-51.
LETTER
TO THE
EDITORS
myo-Inositol Content of the Lens in Hereditary Cataract Mice The free myo-inositol content in the mammalian eye lens is rather high and varies with pathologic states. van Heyningen (1957) reported the decreased mvo-inositol level in the completely opaque lens of the rabbit after X-ray irradiation. Low mvoinositol content, was also observed in lenses of sugar. diabetic and senile cataracts. Studies on the mechanism of developmental and morphological changes in hereditar? cataract have Ijeen performed using the Nakano mouse (Nakano et al., 1960; Iwata and Kinoshita. 1971; Hamai, Fukui and Kuwabara, 1974; Fukui. Gbazawa aml Kinoshita, 1976). We reported here changes of free and lmuntl myo-inositol contmts in lenses of the Nakano cataract mice. _A uralr Nakano mouse was crossed with a BALB/c female. Their litters were mate(l to obtain hon~ozygous mice which developed cataract within a month after birth. Their oRspring were used. BALB/c mice served as normal controls. myo-Inositol was assayed by gas-chromatography as its trimethylsilyl et’her by the method of Yamakawa and Ucda (1964). Gas-chromatographic determination was carried out with R”,, OV-1 on Gas-Chrom P and 5% Ucon LB-550 X on Gas-C’hrom CLH in a glass colunm 3 mm x 2 m. To determine bound myo-inositol in lipids, dried lenses were extracted with chloroform methanol mixture (2 : 1, v/v) according to Folch. Lee and Sloane (1957). The est’ract was once washed wit*h one fifth volume of water to remove free
Age (days) FIG. 1. Concentration of free myo-inositol in the lens of normal (o-0) Each value represents an average of two or three determinations pooled
and cataract (c-9) lenses.
mice.
I20
1.:. \\..\I).\
15T .\I..
sugar atltl polvols. The tlrietl lipid fraction \ws nt~+Jtanolizct1I\-itlt (Fi”,,) iJJl(ltreated with IK AK, tlriwl. ildJICl tritttcth~lsilylatetl. The
COJltf?Jlt
Of
Ill~O-itlOSitO1
it1
th?
hh.
kidney.
tf?StiS
;tll(I
Itl~~t~llil~tl~ll IIC ‘I
hvtht \VilS ;lt il. ~ittll~ilt
level in normal aid cataract mice. In the lens. howt~vcr, thc$ tiiyo-itiositol cotttt~tttN was alqnwximatel,y 5W,, 1on.er in the cataract than in the nortttal tttottw OII ~4; weight I)asis. It was evident t,liat Oh(~hcreditarv tleficiencv afYectot1t,ltt* I(~\YI 01’f’rc~ myo-inositol only in the lens but, not; in ot.lwr-orgarls. I\;-at,a and Kittosltit,;t ( 197I) also oltserved dcficiencv of Nat K ATYaw onlv in the l~~nses of Nakano cataract. IIIIW. Figure l(a) show tler;~lopttrental chat1geh of’ tttvo-inositol _ 1~~~~1 in IJotlt nortttal ;rt~tl cataract, JIJOIJW lenses.The level was the samein I)oth Ictw~sttntjil 10 da!;> :tf%c~r I)irt,tt. Therea,fter. the tnyo-inositol content8 of ttortnal tttonse Irns incrcwecl cotttitluott,sl! to reach ntore than 6 &lenh. In the cataract iJ~01tw lens. howc~vt~l~. i?hc c~lttt~f’tlt increased gratluallv until 30 (tars of age and was then inaittt~aittc~l itt, it htiiitillii;tI value of al)out( I &lrrts. On t’hc: Ilt#h day when Iwth tnicc opettwl t#hc>irt’vt~ atul cataract niicc had transparent8 cbw, thrx lens niyo-inositol l(>wl ~vits sligltt,l;; iowr in the cataract JJ~OIJSCL Ohan in the norrttal. Thus thrl tlly-ill~JShl~ cottttqt of t It,% cataract tttome lens was 60” I, of the JloJ’JJla~ kW?l at f3 \V~xkS (Jf W&y! 1vhCll “pill-tlV;t(l” opacity appeared and only 20” ,, of the normal at 3 months. Whew esprtwe~l on ;I cln weight basis.the myo-inositol content continued to increaseat) it sitttilar rate III) to tttc: age of 50 days in both normal an(l cataract mice with a lag lwriotl of whoub20 cla\-s in the lat,w [Fig. 1(tt)]. Howevcrt it r~w~ainetlat the sattle thrwaftrr in the (~ttt~>trit.ct lens, suggestingthat leveling off to putit]) acbivitv ma\- occur in th(slcrls atj t,ltis agc~. ~OJTeh~i~JJl of rtiyo-inositol level a.11~1 opacity ;,f cataract Icws w-as t’XiJ,ttliJlt?(~ (‘lht)lfs 1). Two litters consisting of tttice with letlses varying itt opacity ant1 OJKS with (.‘l+BitJ’
No. of exp.
1
2
Age (days)
40
34
x0. of lenses
Lens weinht c (w) wet dry
2 4
423 3.1
2
3.2
2 4 4
* Determined macroscopically. - Almost normal, + slight
Degree (right)
2.9
I.21 I.25 1.1” 0.94
++ + i- t-
2.7 24
1.17 0~90
i.+--
cortical
opacity,
of opacity (left)
-t
-k-!-t++
1.7 I.3 14 0.9
-
1.7 9.7
-
-t + marked
+++
opacity
pg of myo-lnositol (/lens) (/rng dry wt.)
and + + + conlplete
140 Ia4
WY!l U.96 143 Cl-SY
opacity.
lenseswere specially selected and divided into four groups. It was remarkablr~ that the lensesfrom these mice, even though transparent, were far lower in u-eight than those of normal mice (5 mg or more) of the sameages. This suggeststhat distur1tattce of lens growth may occur prior to opacification. The clear lens had 1.i pg of ntvc~inositol content corresponding to 70-800,; of the normal. The content was tttarkedly lower in the opaque lenses than in control lenses. It is of interest that the myoinositol content was decreasedwith degree of opacification.
.\IYO-INOSITOL
IX
HEREDITARY
(‘ATARA(“I
121
13otlr lipids and bound myo-inositol content were higher in the cataractous lens thm the normal when expressed as per unit of dry weight (Table II). However, taking account of smaller size and higher water content in the cataract lens. no differelm was found in bound myo-inositol content per lens between cataract anti H~I-I~:L~ mice. It should be pointed out that the amount of hound nlyo-inositol is apparentl,y a small fraction of total mnyo-inositol. In the normd mouse lens free autl Itountl uryo-inositol content was 3.87 and OC ~nml~g wet weight. respectively. In
the cataract lens they were 1.85 and 0.04 +lol, respectively. The ratio tJf IK~LUK~ t,cj myo-inositol was O*52o/o in the normal lens and 2.16” ,, in the cataract lens. Thr Immtl inositol content in the mouse lens was lower by two figures thm that, in rat, tissws reported by Wells, Pittman and Wells (1965). The cataract luoust! lens is sharply contrastStd with the normal one in markedly lowr free inositol cont,cnt. Since phospholipitls are important constituents of t’he nwmhrane syst,em, it is concciva.l)le that t~ht~ membrane system inclutlin g the lens capsule tlicl not, untlrrpo intense conqmitional changes. If there are arl,v defects in the nwnIbmne systcrn, tNlwv we rather functional disorders. frer
KI~FFR~ESCES , , Folch.
J., Let, AI. and Sloane, S. G. H. (195i). -1 simple method for the isolatiorl of total lipids from animal tissues. J. Hid. Chum. 226, 41%319. Hamai, T., .Fukui, H. N. and Kua-abara, ‘I’. (1974). Morphology of hereditary
JSxp. Eye Hes. 18,
at~tl p~lrijicntiorl ~IOIIW
cxt;lrar.t.
537-415.
ran Heyningen, R.. (1957). Mesoinositul in the lens of mammalian eyes. UiocJwu. J. 65, 24 S. Fukui. H. S.. Obazawa, H. and liinoshita. -7. H. (19ili). Lens gron th in the Sakano ~OLISC. Invest. Ophthrrlmol. 15, 422--S. Iweta, S. and Kinoshita. J. H. (19il). !Ucchanism of tlelelopment of hereditary cataract in rrriw. I/west. Ophthdmol. 10, :il)l-1’. Sakano, K., Yamamoto. S.. Kutsukake, C:., Ogawa, H., Sakajima, .I. and Takano. E. (LRtiO). Hereditary cataract in mice. .Jap. J. C’lin. Ophthd. 14, 196-200. Wells, \V. \V., I’ittman, T. A. and IVeIls, H. J. (1965). & uantibtive analysis of myo-inoaitol in rat tissue by gas-liquid chromatography. Annlyt. Hiochem. 10, 450-8. Tamakawa, T. and Veda. S. (1964). C;as-liyuid chromatography of carbohydrates. J~rl/. J. h’.rp.
Jfed. 34, 3-51.