Histological study of the effect of potassium dichromate on the thyroid follicular cells of adult male albino rat and the possible protective role of ascorbic acid (vitamin C)

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Journal of Microscopy and Ultrastructure

jo ur nal homep age: www.els evier .com/ locate / jmau

riginal Article

istological study of the effect of potassium dichromate onhe thyroid follicular cells of adult male albino rat and theossible protective role of ascorbic acid (vitamin C)

eda H. ElBakry, Sadika M. Tawfik ∗

istology Department, Faculty of Medicine, Tanta University, Egypt

r t i c l e i n f o

rticle history:eceived 13 January 2014eceived in revised form 16 April 2014ccepted 22 April 2014vailable online xxx

eywords:istological changesotassium dichromatehyroid glandatscorbic acid

a b s t r a c t

Occupational exposure to toxic heavy metals renders the industrial workers with vari-ous health problems. Many heavy metals act as endocrine disruptors. Chromium salts arecommonly used in industries as asbestos brake lining, cement dust and food additives.Thyroid is a metabolically active important endocrine gland. Little is known about the toxiceffects of chromium on thyroid. So, the aim of this study was evaluation of the histologicalchanges induced by chromium on thyrocytes and the possible protective role of vitamin C.Forty adult male albino rats were divided into four equal groups: control group, vitamin Ctreated group, potassium dichromate treated group and the fourth group co treated withpotassium dichromate and vitamin C. Potassium dichromate was given intraperitonially for2 weeks at a dose 2 mg/kg daily. Vitamin C was given at a dose 120 mg/kg orally daily forthe same period. Specimens were processed for light and electron microscopy. Apoptosiswas evaluated immunohistochemically. Specimens of potassium dichromate group showeddisturbance of the normal architecture of the gland with coalescence and degeneration ofthyroid follicles and desquamated cells in its lumen. Disruption of the apical and basalmembranes of some thyrocytes, flattened hyperchromatic nuclei, dilated RER and swollendegenerated mitochondria were also noted. Immunohistochemically, there were changes
in the immune expression of Bcl2 in the cytoplasm of thyrocytes. Vitamin C supplementedgroup showed partial improvement of the previous changes. So, potassium dichromateinduced structural changes in the thyroid follicular cells that were partially improved byvitamin C supplementation.
© 2014 Saudi Society of Microscopes. Published by Elsevier Ltd. All rights reserved.

. Introduction

Chromium (Cr) is a naturally occurring heavy metalound in rocks, volcanic dust and gases, soils as well as

Please cite this article in press as: ElBakry RH, Tawfik SM. Histthe thyroid follicular cells of adult male albino rat and the possUltrastruct (2014), http://dx.doi.org/10.1016/j.jmau.2014.04.00

lants and animals. Chromium compounds are used inaints, metal finishes, and stainless steel. Increased usagef hexavalent chromium (CrVI) and its improper disposal

∗ Corresponding author. Tel.: +20 01001085481.E-mail address: sadika [email protected] (S.M. Tawfik).

http://dx.doi.org/10.1016/j.jmau.2014.04.003213-879X/© 2014 Saudi Society of Microscopes. Published by Elsevier Ltd. All ri

lead to various health hazards [1,2]. However, chromiumin a small amount is considered an important nutrientresponsible for carbohydrate metabolism [3].

Chromium is present in two forms: trivalent Cr(III)which is poorly transported across membranes and hexa-valent Cr(VI) can readily cross cellular membranes. Insidethe cell, the hexavalent form is reduced to the triva-

ological study of the effect of potassium dichromate onible protective role of ascorbic acid (vitamin C). J Microsc3

lent form. This biotransformation process reduces thetoxicity because the trivalent form does not cross cel-lular membrane rapidly. The trivalent form complexeswith intracellular macromolecules, these toxic compounds

ghts reserved.

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are responsible for many toxic and mutagenic effects ofchromium, not biodegradable and accumulate in the envi-ronment [4,5].

Hexavalent chromium usually linked with oxygen form-ing strong oxidizing agent that is widely known to causeallergic dermatitis as well as toxic and carcinogenic effectsin humans and animals [6]. The fate of chromium in theenvironment is dependent on its oxidation state. Hexa-valent chromium primarily enters the cells and undergoesmetabolic reduction to trivalent chromium, resulting in theformation of reactive oxygen species together with oxida-tive tissue damage and a cascade of cellular events [7].

Occupational exposure to Cr is found among approxi-mately half a million industrial workers worldwide. Also,water contaminated with hexavalent chromium is a world-wide problem, as it is the major route of chromiumexposure for the general population [8,9]. Potassiumdichromate (K2Cr2O7) is a soluble hexavalent chromiumcompound that is widely used in several industries [10].

Ascorbic acid (vitamin C) is as an essential micronutri-ent that performs important metabolic function in human[11]. It also acts as a biological antioxidant by donating anelectron to free radical species, such as tocopherol radical,thereby interrupting the radical chain reaction in biologicalmembranes [12].

Vitamin C has been reported to function as a majorreductant of Cr(VI) in animals and cell culture systems [13].From these points it was important to evaluate the effectof potassium dichromate on the thyroid gland of rats andthe possible protective effect of vitamin C.

2. Materials and methods

The present work was carried out on 40 adult malealbino rats that were divided into four groups each, 10rats. Group one acts as a control group that was subdividedinto two subgroups: A positive control which received intraperitoneal injection of 1 ml distilled water, the solvent ofthe material used in this work. The other subgroup B whichwas the negative subgroup that did not receive any treat-ment through the experiment.

The second group received vitamin C in dose of120 mg/kg [14] orally for 2 weeks. The third group wasthe treated group received potassium dichromate in a doseof 2 mg/kg [15] intra peritoneal per day for 2 weeks. Thefourth group (the protective group) received the same doseof potassium dichromate concomitant with vitamin C inthe same dose like the second one. Potassium dichromate(K2Cr2O7) was supplied by El-Nasr pharmaceutical chemi-cals company ADWIC, Egypt.

At the end of the experiment the rats were anaes-thetized (better with carbon dioxide). Mid line incision wasdone with careful dissection of the neck to identify ster-nohyoid and sternomastoid muscle. The muscles were thenseparated to expose the trachea. Trace the trachea upwardgently until the thyroid glands were visible. It appeared astwo small reddish oval masses on each side of the trachea.


Dissect the glands gently to avoid its injury [16]. The thy-roid glands were collected and divided into two parts. Onepart was processed for light microscope study using PASstain and some sections used for immunohistochemical

PRESS and Ultrastructure xxx (2014) xxx–xxx

methods for detection of apoptosis using BCL2 immuneexpression. The second part was processed for electronmicroscope study. The blood was drawn directly from theleft ventricle through cardiac puncture. It was used forestimation of serum T3, T4 and TSH according to the instruc-tions of their referred methods using Milliplex.Map RatThyroid Hormone TSH panel-2 plex (EMD Millipore, Bil-lerica, MA, USA). The biochemical data were expressed asthe mean ± standard error using ANOVA test (f test) andunpaired Student’s test (Scheffe test).

2.1. Immunohistochemical staining for detection of Bcl2protein (antiapoptotic marker) [17]

Immunohistochemical reaction was performed usingavidin biotin peroxidase technique. The primary antibodywas a rabbit monoclonal antibody) Sigma Laboratories).The universal kit was avidin biotin peroxidase producedby Nova Castra Laboratories Ltd, UK.

Tissue sections were deparaffinized and rehydrated.Slides were incubated in hydrogen peroxide (10%) for10–15 min to inhibit the activity of endogenous peroxi-dase to reduce nonspecific background staining. Antigenretrieval was done by immersing the sections in a pre-heated citrate buffer solution (pH 6) and maintaining heatin a microwave at 2 W for 10–20 min. Sections were left tocool for 20 min at room temperature. Slides were washedfor 2 times in buffer (0.05% sodium azide). Monoclonalantibody Bcl2� Ab-1 (Ms-123-R7) was applied. Slideswere washed for 4 times in buffer (0.05% sodium azide).Biotinylated goat anti-polyvalent was applied. Slides wereincubated for 10 min at room temperature. Slides werewashed for 4 times in buffer (0.05% sodium azide). Chro-mogenic substrate was applied (diaminobenzidine) DABand incubated until desired reaction was achieved. Mayer’shaematoxylin was used as a counter stain [18,19].

The Bcl2 cytoplasmic site of reaction was stained brownand nuclei stained blue. The same method was applied toprepare negative control sections but the primary antibodywas not added. Tonsil was used as positive control tissue[20].

3. Results

3.1. Biochemical results

Potassium dichromate treated rats showed a significantdecrease in the serum T3 and T4 and a significant increase inthe TSH levels when compared with control group and vita-min C treated group. In the protective group, there was asignificant increase in the level of the T3 and T4 and a signif-icant decrease in the serum TSH compared with potassiumdichromate treated group (Tables 1–3).

3.2. Light microscopic results

3.2.1. Control and vitamin C treated groups


Examination of the semi-thin sections obtained fromthe thyroid glands of the control group and vitamin Ctreated showed the normal histological structure. The thy-roid gland was formed of follicles of various sizes, each

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Table 1The mean level of serum tri-iodothyronine (T3) in different studied groups.

T3

GI GII GIII GIV

Mean 112.2 116 81 98±SD 25.6 19.3 15.3 12.7f test 6.325p value 0.006*

Scheffe test

GI & GII GI & GIII GI & GIV GII & GIII GII & GIV GIII & GIV

0.052 NS 0.001* 0.009* 0.001* 0.006* 0.012*

0

20

40

60

80

100

120

mea

n va

lue

GI GII GIII GIV

Histogram s howing the me an lev el ofserum tri -iodothyronine (T3) in different studied groups.

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D, standard deviation; NS, non-significant.* Significant.

ollicle was lined with a single layer of cuboidal epithelialells and filled with homogenous colloid and separated by


apillary beds (Fig. 1).Basal lamina of follicular cells showed moderate PAS

eaction while colloid showed marked reaction witheripheral vacuolization (Fig. 2).

ig. 1. Photomicrographs of a semi-thin section of the thyroid gland of the conlled with homogeneous colloid (F) and separated by capillary beds (C) (Toluidin

Immunohistochemically, there was strong cytoplasmicBCL2 expression in follicular cells (Fig. 3).


3.2.2. Potassium dichromate treated groupSemi-thin sections of the thyroid gland of the potas-

sium dichromate treated rats showed complete loss of the

trol group showing thyroid follicles lined with cuboidal epithelium ande blue, 1000×).

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Table 2The mean level of serum thyroxine (T4) in different studied groups.

T4

GI GII GIII GIV

Mean 9.3 6.96 4.9 4.7±SD 1.63 1.20 0.36 0.63f test 3.265p value 0.042*

Scheffe test

GI & GII GI & GIII GI & GIV GII & GIII GII & GIV GIII & GIV

0.142 NS 0.001* 0.001* 0.014* 0.010* 0.020*

0

2

4

6

8

10

mea

n va

lue

GI GII GIII GIV

Histogram showing the mean level of serum thyroxine (T4) in different studied groups.

SD, standard deviation; NS, non-significant.* Significant.

ing stropheral v

Fig. 2. Photomicrographs of the thyroid gland of the control group showreaction in the basal lamina. Notice, in (A) there are follicles with no peri

normal architecture of the thyroid gland. Disruption ofthe basal laminae of some follicles with their coalescencewas detected (Fig. 4). Some follicles appeared distendedwith colloid lined with squamous epithelium, others werecompletely collapsed. Some follicles were degenerated andfilled with exfoliated cells (Fig. 5). Disruption of the apicalmembranes of some thyrocytes with desquamated cyto-


plasm and nuclei inside the follicles was observed. Somethyrocytes appeared tall columnar with apical pseudo-podia (Fig. 6). Vacuolization of the cytoplasm of some

ng PAS reaction in the colloid with peripheral vacuolation and moderateacuolation (PAS, 400×).

thyrocytes, variable densities of colloid staining and widen-ing of the interfollicular space appeared in some sections(Fig. 7).

Basal lamina of follicular cells showed a weak PASreaction while colloid showed a moderate reaction withabnormal pattern of vacuolization and colloid retraction insome follicles (Fig. 8).


Immunohistochemically, there was negative cytoplas-mic BCL2 immunoexpression in the majority of follicularcells (Fig. 9).

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Table 3The mean level of serum thyroid stimulating hormone (TSH) in different studied groups.

TSH

GI GII GIII GIV

Mean 0.01 0.01 0.12 0.01±SD 0.01 0.01 0.02 0.01f test 1.362p value 0.069*

Scheffe test

GI & GII GI & GIII GI & GIV GII & GIII GII & GIV

1.00 NS 0.001* 1.00 NS 0.001* 1.00 NS

0

0.02

0.04

0.06

0.08

0.1

0.12

mea

n va

lue

GI GII GIII GIV

Histog ram s howing the mean level of serum thyroid stimulating hormone (TSH) in different studied groups.

SD, standard deviation; NS, non-significant.* Significant.

F ing stronc

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ig. 3. Photomicrographs of the thyroid gland of the control group showells (BCL2 immunoexpression, 400×, 1000×).

.2.3. Protective group (potassium dichromate + vitamin)

Examination of semi-thin sections obtained from thehyroid glands of rats of the protective group revealedmprovement in the histological changes detected in the


revious group. The thyroid gland started to regain its nor-al architecture. The densities of colloid staining were

lso variable. However, some follicles still disrupted withesquamated cells in the lumens. The thyrocytes appeared

g BCL2 immunoexpression in the cytoplasm of the majority of follicular

less vacuolated. The interfollicular spaces appeared nar-rower than the treated group (Figs. 10 and 11).

The basal lamina of the thyroid follicles showed appar-ent increase in the PAS reaction but less than the controlgroup. The colloid showed variable degrees of PAS reaction


and still vacuolated (Fig. 12).Immunohistochemically, there was apparent increase

in the BCL2 expression in the cytoplasm of the thyroid fol-licular cells but less than the control group (Fig. 13).

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Fig. 4. A photomicrograph of a semi-thin section of the thyroid gland ofpotassium dichromate treated group showing loss of the normal architec-ture of the thyroid gland (F), the basal lamina of the follicles are disrupted(arrows) with adherence of the follicles (Toluidine blue, 400×).

Fig. 5. Photomicrographs of a semi-thin section of the thyroid gland of potassiuwith colloid (F), others appear collapsed (C). (B) Higher magnification of A, showdegenerated (D) with desquamated cells in the lumen (arrows) (Toluidine blue, 4

Fig. 6. Photomicrographs of a semi-thin section of the thyroid gland of the same gwith the presence of the cytoplasmic content in the lumen (S). Notice vacuolatio(B) Apical pseudopodia (thick arrows) of some thyrocytes while the others have v


3.3. Electron microscopic study

3.3.1. Control and vitamin C treated groupsEM examination of ultra-thin sections of the thyroid

glands of rats of the control and vitamin C treated groupsshowing the same ultrastructure picture of normal thy-roid follicular cells. The thyrocytes appeared with roundedeuchromatic nuclei and numerous short apical microvilli.Their cytoplasm contained RER, abundant mitochondria,prominent Golgi apparatuses, SER, lysosomes, abundantapical vesicles containing colloid and electron dense gran-ules (Fig. 14).

3.3.2. Potassium dichromate treated groupEM examination of ultra-thin sections of the thyroid

glands of this group confirmed the light microscopic exam-ination. Some of the thyrocytes were squamous with flathyperchromatic nuclei, others were pyramidal or prismlike. Stratification of the lining epithelium of some fol-


licles and desquamated cells appeared in the follicularlumens (Fig. 15). The cytoplasm showed variable sizedvacuoles with dilated RER with disruption of the api-cal membrane of some thyrocytes and exfoliated cells

m dichromate treated group showing: (A) some follicles are distendeding some of thyroid follicles appeared collapsed (C), while the others are00×, 1000×).

roup showing (A) disruption of the apical membranes of some thyrocytesn of the cytoplasm of the other thyrocytes in the same follicle (arrows).acuolated cytoplasm (thin arrows) (Toluidine blue, 1000×).

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Fig. 7. Photomicrographs of a semi-thin section of the thyroid gland of the same group showing (A) widening of the interfollicular space (stars) and variabledensities of the colloid staining (v). (B) Mast cells in the interfollicular space (arrows) (Toluidine blue, 1000×).

Fig. 8. Photomicrographs of the thyroid gland of potassium dichromate treated gthe basal lamina with (A) absence of peripheral vacuolation and (B) abnormal pat

Fig. 9. A photomicrograph of the thyroid gland of potassium dichromatetreated group showing negative immune expression of BCL2 in the cyto-p

wtcf(

sible protective role of vitamin C. The study revealed theeffects of chromium compound on the thyroid gland which

lasm of the majority of follicular cells (BCL2 immunoexpression, 1000×).

ere present inside the lumens (Fig. 16). In some sec-ions there is rarefaction of the cytoplasm with loss of theytoplasmic organelles swollen degenerated mitochondria,


ragmented RER and the presence of large sized vacuoleFig. 17).

roup showing moderate PAS reaction in the colloid and weak reaction intern of vacuolization and colloid retraction (PAS, 400×).

3.3.3. Protective group (potassium dichromate + vitaminC)

The follicular cells of this group appeared more or less asthe control group. However, some thyrocytes still have vac-uolated cytoplasm, dilated RER and hyperchromatic nuclei(Figs. 18 and 19).

4. Discussion

Hexavalent chromium (CrVI) has been widely used inindustries throughout the world. Increased its usage andatmospheric emission of CrVI from catalytic convertersof automobiles, and its improper disposal cause varioushealth hazards [2].

Today, it is known that there is a close relationshipbetween workers health and the amounts of industrial pol-lution provoked by industries manufacturing chromiumcontaining materials. So, numerous and different in vitroand/or in vivo studies on chromium have been undertakeneither in animals or in human [21].

The present work studied the effect of potassiumdichromate on the thyroid gland structure and the pos-


appeared in the form of irregularity in size and shapeof the follicles. Some of them were lined with squamous

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Fig. 10. Photomicrographs of a semi-thin section of the thyroid gland of the protective group showing: (A) restoration of the normal architecture of thethyroid gland (F). (B) Disruption of the basal lamina of the thyroid follicles (arrows) (Toluidine blue, 400×).

Fig. 11. Photographs of a semi-thin section of the thyroid gland of the same group showing (A) the thyroid follicles appear more or less as the control (F)(Toluidine blue, 400×). (B) Higher magnification of A showing vacuolization (arrows) of the cytoplasm of some thyrocytes (Toluidine blue, 1000×).

showinear wit

Fig. 12. Photomicrographs of the thyroid gland of the protective grouplamina, some follicles showing peripheral vacuolization, while others app

epithelium, others lined with cubical and the third groupof follicles were lined with tall columnar cells with apicalpseudopodia. Interfollicular spaces were widened either asa result of collapse of some follicles or due to necrosis of theothers leaving empty spaces. The cytoplasm of some follic-ular cells appeared vacuolated and even desquamation of


some cells into the lumens were seen in the majority ofsections.

g strong PAS reaction in the colloid and moderate reaction in the basalh central vacuolization, the rest appear inactive (PAS, 400×).

Other work reported that, administration of sodiumdichromate in the drinking water to rats and mice resultedin focal ulceration, hyperplasia, and metaplasia in the glan-dular stomach and hyperplasia of the duodenum [22]. Also,histological analysis of the effect of chromium on malereproductive system revealed pronounced morphological


alterations with enlarged intracellular spaces and tissueloosening [15].

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The present work revealed increase in the interfollic-lar spaces and disruption of the basal lamina of folliclesnd these findings were coincided with other studies [23].he latter reported decrease in the size of follicles andidening of the interfollicular spaces and attributed thisidening to disruption of the connective tissue. At the end

hey concluded that Cr(VI) is potentially toxic to the thy-oid gland. Also, another studies found the same resultsith irregularity and shrunken of some follicular cells

ncrease in the number of disorganized and collapsed fol-icles. Hyperplasia of follicular cells and nuclei appearedval, rounded, irregular and even shrunken. They addedhat disruption of the apical and basal laminae were seen24].

Disorganization of the thyroid follicles and cytoar-hitecture indicate high level of toxicity caused by thehromium. TSH is responsible for the morphologicalppearance of thyroid follicles and the synthesis and secre-ion of thyroid hormones leading to hypertrophy andyperplasia of the follicular cell. One of the early responsesf TSH-stimulated thyroid follicular cells is engulfment ofolloid material from the follicular lumen into the apicalytoplasm of thyrocytes, in the form of membrane-boundolloid droplets. Administration of TSH to rats pretreatedith thyroxin resulted in the formation of numerous pseu-opods on the apical surface of thyrocytes, followed by theppearance of colloid droplets, at first in the apical andater on in the deeper parts of the cytoplasm. It was alsotated that the percentage of follicular cells containing col-oid droplets and the number of droplets in cells graduallyncreased with the increase of TSH dose [23,25].

However, irregularity in the size of follicles in some ani-


als may be normal, as the size distribution pattern ofhyroid follicles is different between the peripheral andentral gland regions. In rats, the larger thyroid gland fol-icles are mostly located at the periphery of the thyroid

ig. 14. (A) Ultrathin section of the thyroid gland of the control group showing

ER (r), scattered electron dense vesicles (arrows). (B) Higher magnification of (Aesicles (arrows).

Fig. 13. A photomicrograph of the thyroid gland of the protective groupshowing positive cytoplasmic reaction for BCL2 (BCL2 immunoexpression,1000×).

lobes stated that the large follicles at periphery mainlyserve as a pool of old hormone, whereas the smaller, cen-trally distributed ones are responsible for thyroid hormonesecretion [26,27].

Variation in the nuclear shape and size was characteris-tic finding in this work and explained by some authors thatincreased nuclear area is often observed in compounds thatblock cell cycle and induce DNA damage [28].

Bcl-2 is an apoptosis-related molecule that was shownto be down-regulated during the early events leading toprogrammed cell death and its protein might be as aninhibiting molecule and play an important role in the bal-


ance between apoptosis promotion and inhibition [29,30].The detection of apoptosis in the follicular cells in

this study which was explained in many other studies[31,32]. They stated that CrVI induced DNA fragmentation,

follicular cells with apical microvilli containing euchromatic nuclei (N),) showing parts of two follicular cells, containing RER (r), electron dense

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Fig. 15. (A) Ultrathin section of the thyroid gland of the potassium dichromate treated group showing two thyrocytes one of them is squamous withflat hyperchromatic nucleus (N1) and the other is prism like with irregular hyperchromatic nucleus (N2). (B) Stratification of the lining epithelium (S),vacuolation of the cytoplasm (V) and exfoliated cells in the lumen (curved arrows).

ate trea) with thlls (arro

Fig. 16. Ultrathin section of the thyroid gland of the potassium dichrom(curved arrow). (B) Higher magnification of (A) showing dilated RER (Rcomplete loss of the microvilli on the apical membrane of the follicular ce

increased apoptosis, increased cytochrome c release fromthe mitochondria to cytosol, down regulated anti-apoptoticBcl-2 and other mediators; upregulated pro-apoptotic[2]. Also it was documented that potassium dichromate-induced apoptosis and oxidative stress in the hepatocytesof Wistar rats.


Some authors explained in details the mechanism bywhich the Cr(VI) lead to apoptosis. They hypotheses thatsoluble Cr(VI) is metabolized within cells by reductive

ted group showing (A) dilated RER (R) and exfoliated cells in the lumene presence of various sized electron dense bodies (arrows). Notice thew head).

agents including ascorbic acid, glutathione and cysteineand a diverse range of genetic lesions are generated. Someforms of Cr damage present physical barriers to DNA repli-cation/transcription and promote a terminal cell fate suchas apoptosis or terminal growth arrest. Other DNA lesionsare potentially pre-mutagenic and lead to DNA damage and


cell cycle arrest [33–35].The electron microscopic results confirmed the results

obtained by the light microscope. Some follicles were

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Fig. 17. Ultrathin section of the thyroid gland of the potassium dichromate treated group showing (A) a thyrocyte with large sized vacuole (V), rarifiedcytoplasm (star), swollen degenerated mitochondria (curved arrows), fragmented RER (r) and flat hyperchromatic nucleus (N). (B) Thyrocytes with irregularshaped nucleus (N) rarified cytoplasm (star), fragmented RER (r) as well as partial loss of the apical microvilli (arrows).

F showino (V). (B) A

llwwtitsra

ig. 18. (A) Ultrathin section of the thyroid gland of the protective groupf the apical membrane (curved arrows) and small cytoplasmic vacuoles

ined with low prismatic, cuboidal or flattened epithe-ium with apparent decrease in epithelial height. That

as attributed to hypoactivity of the thyroid gland. Thisas confirmed by biochemical changes in this work in

he form of decrease in T3 and T4 level and increasen the TSH level [36]. Others showed stratification of


heir lining epithelium. The nuclei of thyrocytes appearedhrunked hyperchromatic nuclei, fragmented and dilatedER, swollen mitochondria with degenerated cristae,ccumulation of colloid vesicles inside the thyrocytes, even

g (A) thyrocytes appear more or less as the control except for disruption thyrocyte with intact mitochondria (M), RER (R) appear as the control.

complete loss of all the cytoplasmic organelles. Similarresults observed previously in the form of disrupted basallaminae of the follicles, regressed nuclei and disrupted cellorganelles [24]. Also, follicular hyperplasia may be con-nected with a cascade of cellular events involving oxidativestress, genomic DNA damage, and modulation of apop-

53


totic regulatory gene (P ) after exposure to chromium[37]

The mechanism of chromium toxicity was reported tobe associated with mitochondrial and lysosomal injury by

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p showilated RE

Fig. 19. (A) Ultrathin section of the thyroid gland of the protective groushaped nuclei (N), rarified cytoplasm (white stars). (B) A thyrocyte with d(arrows) with loss of the apical microvilli (curved arrow).

biologically Cr(VI) reactive intermediates and reactive oxy-gen species [4]. The results demonstrated that potassiumdichromate was highly cytotoxic to cells, and its cytotox-icity seems to be mediated by oxidative stress and DNAdamage [7]. The results indicated that administration ofCr(VI) had caused a significant increase of reactive oxygenspecies (ROS) level generated during reduction of Cr(VI)[38,39].

Previous studies both in vitro and in vivo have demon-strated that chromium(VI) induces an oxidative stressthrough enhanced production of reactive oxygen species(ROS) leading to genomic DNA damage and oxidativedeterioration of lipids and proteins. A cascade of cellularevents occur following chromium (VI)-induced oxidativestress including enhanced production of superoxide anionand hydroxyl radicals, increased lipid peroxidation andgenomic DNA fragmentation, modulation of intracellularoxidized states, activation of protein kinase C, apoptoticcell death and altered gene expression [40–42].

In a recent study demonstrated that catalase is aclassical oxidative biomarker and is the most abundantin peroxisomes, where oxidative stress most frequentlyoccurs [43,44]. The increase in these enzyme activities sug-gests a response toward increased ROS generation [45,46].Previously, it was reported that Cr(VI) induced thyroidtoxicity through increasing cellular oxidative stress anddecreasing the activity of antioxidants [47].

Some authors explained that Cr(VI) accumulates inthe pituitary and hypothalamus resulting in apopto-sis evidenced by nuclear fragmentation and caspase 3activation. Their results indicate that the anterior pitu-


itary gland can be a target of Cr(VI) toxicity in vivoand in vitro, thus producing a negative impact on thehypothalamic–pituitary-axis and affecting the normalendocrine function [9,48].

ing part of thyroid follicle in which the thyrocytes appear with variableR (R) and irregular hyperchromatic nuclei (N) and electron dense bodies

Some researches observed hypertrophic follicular cellscontain dilated r-ER and colloid droplets. Follicles werelined by tall columnar cells and vacuolated mitochondriawith disrupted cristae. They explained their results dueto Goitrogenic effects of a soybean diet which result inincreased thyroid stimulating hormone leading to hyper-plasia in thyroid gland [49].

As regard to the thyroid hormone level there wasdecrease in the level of free T3 and T4. The protectivegroup revealed more or less normal value near to that ofthe control group. The same results were documented inother works which showed significantly increased TSH anddecreased FT3 and FT4 concentrations in protective groupserum TSH, FT3 and FT4 concentrations neared control [14].It was reported that Cr toxicity led quite possibly to a stateof hypothyroidism as indicated by a significant increase inthe serum TSH and a decrease in the serum FT3 and FT4concentrations [50].

Vitamin C administration in this work showed someimprovement in the thyroid gland function and structure.As, the level of the thyroid hormones became near to thecontrol one. Also, the thyroid gland regained its normal cel-lular architecture and appeared more or less near to thecontrol.

Previous studies detected that early and repeated highintravenous doses of vitamin C was the therapy of choicefor Cr(VI) poisoning [51]. Also the results of [52] showedthat vitamin C had antimutagenic effect against chromiumcompound.

Also it was found that administration of exogenousascorbic acid has been used in the treatment of systemic


chromium poisoning and chromium dermatitis as itenhances the extracellular reduction of chromium(VI)[53]. Ascorbic acid influenced Cr(VI) toxicity either byacting as a reducing agent, decreasing Cr(V) persistence or

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ARTICLEMAU-32; No. of Pages 14

R.H. ElBakry, S.M. Tawfik / Journal of Mi

y acting as an antioxidant, decreasing intracellular super-xide anion and hydrogen peroxide formation and also itcts by reducing free radicals formed during reduction ofr(VI) to Cr(III) [54].

Many works concluded that co-administration of vit and E may, significantly diminish the toxic effects ofexavalent chromium on lung [55]. Simultaneous admin-

stration of vitamin C significantly prevented the increasen lipid peroxidation and enhanced the antioxidant sta-us. These results proved the protective effect of vitamin

against the Cr(VI) exposure-induced toxicity and testhe significance of antioxidants in diet [3]. The functionalhanges cannot be completely replenished by the ascorbiccid supplementation in response to chromium exposure56]. However, other works proved that the ascorbic acid

ay have the potential to protect thyroid gland fromhromium toxicity [14].

. Conclusion

It could be concluded that potassium dichromate hadoxic effects on the thyroid follicular cells, which partiallymproved with vitamin C co-treatment. So, it is recom-

ended to give vitamin C as a protective agent againsthromium toxicity in people who are susceptible for con-inuous exposure.

onflict of interest

None declared.

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Documents

Histological study of the effect of potassium dichromate on the thyroid follicular cells of adult male albino rat and the possible protective role of ascorbic acid (vitamin C)